Cellular features of localized microenvironments in human meniscal degeneration: a single-cell transcriptomic study
- Department of Orthopedics, Orthopedic Research Institute, West China Hospital, Sichuan University, China
- MOE Key Lab of Bioinformatics, Bioinformatics Division, BNRIST and Department of Automation, Tsinghua University, China
- School of Life Sciences and School of Medicine, Center for Synthetic and Systems Biology, Tsinghua University, China
Figures
Figure 1 with 2 supplements
Single-cell RNA-seq reveals major cell classes in human menisci.
(A) Photographs of typical normal and degenerative meniscus specimens. (B) The overall workflow of the single-cell sequencing. Inner and outer meniscal parts are collected separately from patients …
Figure 1—figure supplement 1
Quality control metrics for single-cell RNA sequencing.
(A) Jittered violin plots show the number of UMI counts observed per cell grouped by the sample. (B) Jittered violin plots show the number of features observed per cell grouped by the sample. (A) …
Figure 1—figure supplement 2
Signature genes of the large cell classes.
(A, B) The cluster marker genes expression levels are shown on the uniform manifold approximation and projection (UMAP) plots with colors. Inner samples: up; outer samples: down.
Figure 2 with 4 supplements
Identification of chondrocyte and PCL subclusters in human meniscus.
(A) Uniform manifold approximation and projection (UMAP) visualization of the chondrocyte and PCL class cells. (B, C) UMAP visualization of the distribution of chondrocytes at different anatomical …
Figure 2—figure supplement 1
Label agreements of the chondrocyte and pericyte-like cells across samples.
(A) Pearson correlations of chondrocyte/PCL subclusters’ expression across all meniscus specimens. (B) The expressions of subcluster signatures genes across all meniscus specimens. The two heatmaps …
Figure 2—figure supplement 2
Gene ontology enrichment of chondrocyte and PCL subcluster markers.
(A–G) Pathway enrichment analysis of seven subpopulations of chondrocytes and pericyte-like cells. The gene set overrepresentation enrichment analysis (GSEA) enrichment was performed using gene set …
Figure 2—figure supplement 3
Regulons derived by pySCENIC.
(A) Binarized activity scores of GRN modules in chondrocytes and pericyte-like cells. Each row represents a GRN module, while each column represents a cell. The activation status of the module in …
Figure 2—figure supplement 4
Public chondrocyte datasets reanalysis (I).
(A) GSE133449’s normal group marker genes of fibrochondrocytes (FC), regulatory chondrocytes (RegC), prehypertrophic chondrocytes (PreHTC), proliferating fibrochondrocytes (ProFC), cartilage …
Figure 3 with 7 supplements
Degeneration molecular patterns in chondrocytes and PCLs.
(A) A volcano plot shows differentially expressed genes with high fold-change values. The comparison was made between (1) degenerated chondrocytes/PCL cells and (2) normal chondrocytes/PCL cells. (B)…
Figure 3—figure supplement 1
Expression levels of selected gene sets.
(A) Expression of extracellular matrix (ECM) decomposition-related genes and anti-decomposition-related genes in chondrocyte subpopulations. (B) Expression of genes associated with angiogenesis and …
Figure 3—figure supplement 2
Public chondrocyte datasets reanalysis (II).
(A) Differentially expressed genes in osteoarthritis (OA). The heatmap shows the differential gene expression of articular cartilage in normal and OA states by reanalyzing public data (GSE114007). (B…
Figure 3—figure supplement 3
Comprehensive integration of public data and in-house data.
(A) Uniform manifold approximation and projection (UMAP) of in-house data as an integration reference. (B) UMAP of public data (GSE133449) after integration. Coordinates and reference labels were …
Figure 3—figure supplement 4
Finding differentially expressed genes (DEGs) in total chondrocytes and PCLs (degenerated vs. normal).
The heatmap shows the z-score-scaled expressions of DEGs in the degeneration group and the normal group. The DEG analysis was conducted on the total chondrocytes and total pericyte-like cells. Cell …
Figure 3—figure supplement 5
Interpreting differentially expressed genes (DEGs) in total chondrocytes and PCLs (degenerated vs. normal).
(A) Lists of selected top DEGs with absolute fold changes >2. (B, C) Enrichment analysis of DEGs in (A). The enrichment was performed using gene set overrepresentation analysis using clusterProfiler.
Figure 3—figure supplement 6
Finding composition changes and differentially expressed genes (DEGs) in individual chondrocyte and PCL clusters (degenerated vs. normal).
(A) Changes in the composition of the seven chondrocyte subclusters of a meniscus in different statuses and different samples. Left: summarized changes, error bars stand for standard deviations. …
Figure 3—figure supplement 7
Shifts of cell type compositions and inflammation states.
(A, B) Kernel density estimation of data points in normal inner/outer and degenerated inner/outer samples. Color darkness represents the kernel density estimation of the data points. Isoclines are …
Figure 4
Immunofluorescent staining of human menisci demonstrating resident cell subtypes.
Representative immunofluorescence staining images. Color settings: MMP14 for Ch.2 cells (orange), SERPINA1 for Ch.1 cells (magenta), ACTA2 for PCL cells (white), CDON for Ch.3 cells (cyan), CYP1B1 …
Figure 5 with 2 supplements
Immune, endothelial cells, and their crosstalk with chondrocytes.
(A) Uniform manifold approximation and projection (UMAP) visualization of the immune cell types in the meniscus. (B) UMAP visualization of the statuses of samples (degenerated/normal). (C) …
Figure 5—figure supplement 1
Immune cell signature genes.
(A) Signature genes that were used to define the immune cell subpopulations. (B) Finding DEGs in macrophages/monocytes (normal vs. degenerated). (C) Finding DEGs in T cells (normal vs. degenerated). …
Figure 5—figure supplement 2
Ligand–receptor pairs that were up-/downregulated in degeneration.
(A–L) The heatmaps reported ligand–receptor pairs that have significant differences across all sender and receiver cell types.
Figure 6
A schematic diagram of the microenvironment changes between the normal and the degenerated meniscus.
The left side visualizes the homeostasis meniscus, where Ch.1 is the dominant chondrocyte population. In this situation, the extracellular matrix (ECM) decomposition and synthesis reach a dynamic …
Additional files
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Supplementary file 1
Donor and sample information.
A table contains the descriptions of sample names, cell counts, cell viabilities, sample volumes, estimated numbers of cells loaded, donor states, donor genders, anatomical locations, estimated numbers of cells after filtering, body mass index (BMI) of donor, and ages of donor.
- https://cdn.elifesciences.org/articles/79585/elife-79585-supp1-v1.xlsx
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Supplementary file 2
Differentially expressed genes of each cluster.
A list of differentially expressed genes of each cluster. The table contains gene symbols, corresponding clusters, average log2 fold-change values, percents of cells expressing this gene in groups 1 and 2, p values, and adjusted p-values.
- https://cdn.elifesciences.org/articles/79585/elife-79585-supp2-v1.xlsx
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MDAR checklist
- https://cdn.elifesciences.org/articles/79585/elife-79585-mdarchecklist1-v1.docx
