Figure 1. A pre-enrichment strategy to... | Rockefeller University Press

Figure 1.

$A pre-enrichment strategy to examine the heterogeneity of pDCs in SSc patients at the single-cell level. (A) Flow chart describing the sample preparation, library construction, and data processing. Frozen PBMCs from four HDs and four diffuse cutaneous SSc patients were sorted to obtain enriched pDCs and T cell–depleted PBMCs fractions, the two fractions were then remerged at 1:2 ratio before loading to 10× single-cell microfluidic chips. After library construction and sequencing, a merged dataset with cells was obtained and batch-corrected using FastMNN in a base of Seurat pipeline. Doublets were removed manually, and the data were visualized by UMAP plots. (B) Cluster annotation of PBMCs. The dot plot represents expression values of selected genes (x axis) across each cluster (y axis). The color intensity indicates the scaled average expression within expressing cells, where red indicates high expression and blue shows low expression. The dot size represents the percentage of cells expressing the marker genes. (C) The UMAP visualization of SCs of merged cells from four healthy and four SSc patients' PBMCs, the putative identity of each cluster was assigned on the basis of B. (D) Comparison of the cell distribution of HDs (blue, n = 13,661 cells) and SSc patients (red, n = 17,714 cells) by UMAP plot. (E) The ratio of the cell number of healthy donors (n = 4) and SSc patients (n = 4) in each SC is shown by a bar plot. (F) Bar plot showing the comparison of the distribution of PBMC SCs between HDs (n = 4) and SSc patients (n = 4). (G) Bar plots highlighting cell abundances of different subtypes of PBMCs for healthy donors (n = 4) and SSc patients (n = 4). (H) Upset plot showing the intersections of the up-regulated (red, log2FC > 0.25, P_adj < 0.05) and down-regulated (blue, log2FC < −0.25, P_adj < 0.05) in different cell types of PBMCs, Wilcoxon rank sum test was used for DEG identification. Statistical significance in G was evaluated using Mann–Whitney U test, and only comparisons that are significant are shown. *P < 0.05.$

A pre-enrichment strategy to examine the heterogeneity of pDCs in SSc patients at the single-cell level. (A) Flow chart describing the sample preparation, library construction, and data processing. Frozen PBMCs from four HDs and four diffuse cutaneous SSc patients were sorted to obtain enriched pDCs and T cell–depleted PBMCs fractions, the two fractions were then remerged at 1:2 ratio before loading to 10× single-cell microfluidic chips. After library construction and sequencing, a merged dataset with cells was obtained and batch-corrected using FastMNN in a base of Seurat pipeline. Doublets were removed manually, and the data were visualized by UMAP plots. (B) Cluster annotation of PBMCs. The dot plot represents expression values of selected genes (x axis) across each cluster (y axis). The color intensity indicates the scaled average expression within expressing cells, where red indicates high expression and blue shows low expression. The dot size represents the percentage of cells expressing the marker genes. (C) The UMAP visualization of SCs of merged cells from four healthy and four SSc patients' PBMCs, the putative identity of each cluster was assigned on the basis of B. (D) Comparison of the cell distribution of HDs (blue, n = 13,661 cells) and SSc patients (red, n = 17,714 cells) by UMAP plot. (E) The ratio of the cell number of healthy donors (n = 4) and SSc patients (n = 4) in each SC is shown by a bar plot. (F) Bar plot showing the comparison of the distribution of PBMC SCs between HDs (n = 4) and SSc patients (n = 4). (G) Bar plots highlighting cell abundances of different subtypes of PBMCs for healthy donors (n = 4) and SSc patients (n = 4). (H) Upset plot showing the intersections of the up-regulated (red, log₂FC > 0.25, P_adj < 0.05) and down-regulated (blue, log₂FC < −0.25, P_adj < 0.05) in different cell types of PBMCs, Wilcoxon rank sum test was used for DEG identification. Statistical significance in G was evaluated using Mann–Whitney U test, and only comparisons that are significant are shown. *P < 0.05.

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