Article Figures & Data
Figures
- Figure 1 Cell Types in CSF of Patients With MS or Other Diagnoses
(A) tSNE plot of cells from CSF. Data were clustered using k-means, and the clusters colored as shown in the key on the right of the figure, in a fashion exactly analogous to figure e-1, links.lww.com/NXI/A386. The characteristics of these clusters are shown in heatmap (C). The tSNE plot on the left shows data from cells from CSF of patients with non-MS diagnoses, the figure in the middle shows data from patients with MS, and the figure on the right shows data from patients with MS treated with rituximab. The red arrow in the bottom right corner of the middle tSNE plot indicates a cluster that is more abundant in CSF from patients with MS than in controls (see below). For this analysis, cells from CSF were clustered and analyzed independently, unlike in figure 1, enabling the detection of clusters that are specific to CSF, and too small to see against the background of the much larger dataset from blood. (B) Areas of the tSNE plots in (A) have been matched to the broad cell types myeloid, NK cells, T cells, and B cells. (C) Heatmap showing, for each of the clusters generated by k-means, the average intensity of each marker across all cells in the cluster. The colored bars at the top of the heatmap match the colors used in the tSNE plots in (A), and clusters corresponding to well-established cell types are shown on the right. Higher average expression of each marker is indicated with a red-brown color, and lower expression in blue.
- Figure 2 Relative Abundances of Cell Types in CSF of Patients With MS or Other Diagnoses
For each donor, the relative abundance of the cluster (% of total cells) was plotted for each cluster described in figure 1. Cluster identity numbers are at the top of each boxplot, and the colors of the bars at the top of each boxplot match the colors used to mark the clusters in figure 1. Within each scatter column, each filled circle represents 1 donor. The diameter of the circles reflects the absolute size of the cluster in that donor. The 35 donors are separated into 3 groups: donors with other diagnoses than MS (control, light gray boxes on the left of each boxplot); donors diagnosed with MS, but not treated with CD20-depleting antibodies (dark gray box in middle); and donors diagnosed with MS and treated with rituximab (gray box on right). Data from 2 rituximab-treated patients are shown for interest without statistical inference. The asterisks mark the cluster in which there was a statistically significant difference in abundance between control and MS (generalized linear mixed model; **p = 0.0012, FDR = 0.0192). FDR = false discovery rate.
- Figure 3 Volcano Plot of Soluble Markers in CSF From Donors With MS or Non-MS Diagnoses
The vertical axis is the log10 of the p value for the comparison. The horizontal axis is the log2 of the fold change. Names of analytes with FDR of less than 0.05 are printed beside the point. Red points correspond to analytes that are significantly more abundant in CSF from patients with MS, and blue points to analytes that are less abundant in MS. Analytes discussed in the text are indicated with arrows, whose directions and colors are explained on the right of the figure. FDR = false discovery rate.
- Figure 4 Principal Component Analysis of Soluble Molecules Measured by PEA
(A) The contribution of each of the soluble molecules to the first 2 principal components PC1 and PC2 is shown by plotting each as a vector. For example, CXCL13 makes a large contribution to PC1, whereas HMOX1 mostly contributes to PC2. For clarity, only the 60 most strongly discriminating molecules are labeled. The broken green arrow pointing down and left indicates a hypothetical axis of neural plasticity, inferred from the preponderance of molecules involved in neural development, axonal signaling, and so on. The red arrow pointing down and right indicates a hypothetical axis of disease activity, inferred from the preponderance of inflammatory molecules, and the presence in this quadrant of early disease stages (see plot B). Candidate MS-associated, plasticity-suppressive chemokines CXCL1 and CXCL8 (see Discussion) are marked with green asterisks. (B) PCA sample scores plot, where each point represents one donor, and the color and shape of each point indicate the diagnosis, disease stage, and treatment of the donor, as shown in the legend below the plot. Green and red arrows are identical in meaning to (A). PEA = proximal extension assay.
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