Inflammation of the choroid plexus in progressive multiple sclerosis: accumulation of granulocytes and T cells

Abstract

The choroid plexus (CP) is strategically located between the peripheral blood and the cerebrospinal fluid, and is involved in the regulation of central nervous system (CNS) homeostasis. In multiple sclerosis (MS), demyelination and inflammation occur in the CNS. While experimental animal models of MS pointed to the CP as a key route for immune cell invasion of the CNS, little is known about the distribution of immune cells in the human CP during progressive phases of MS. Here, we use immunohistochemistry and confocal microscopy to explore the main immune cell populations in the CP of progressive MS patients and non-neuroinflammatory controls, in terms of abundance and location within the distinct CP compartments. We show for the first time that the CP stromal density of granulocytes and CD8+ T cells is higher in progressive MS patients compared to controls. In line with previous studies, the CP of both controls and progressive MS patients contains relatively high numbers of macrophages and dendritic cells. Moreover, we found virtually no B cells or plasma cells in the CP. MHCII+ antigen-presenting cells were often found in close proximity to T cells, suggesting constitutive CNS immune monitoring functions of the CP. Together, our data highlights the role of the CP in immune homeostasis and indicates the occurrence of mild inflammatory processes in the CP of progressive MS patients. However, our findings suggest that the CP is only marginally involved in immune cell migration into the CNS in chronic MS.

Keywords: Blood-CSF barrier; Choroid plexus; Granulocytes; Immune cells; Progressive MS; T cells.

Fig. 1

Density of macrophages and DCs is similar in the CP of progressive MS patients and controls. a Example of the compartmentalization of the CP tissue. Using the basement membrane as guidance (collagen IV, in white), we discern the stromal and epithelium-associated cells (which together are the CP cells), and exclude the circulating cells located in the vessels. b Maximum projection of an image of a CP immunolabeled with Iba1, MHCII and collagen IV. Right panel shows a higher magnification of the image. White arrowheads point to one MHCII+ and one MHCII- macrophage (Iba1+) in the CP stroma; white arrow points to a dendritic cell (DC; Iba1- MHCII+) associated to the epithelium. c Density of CP macrophages (MHCII+ and MHCII-) in control and progressive MS (Wilcoxon rank sum test with continuity correction). d Density of CP macrophages in the different CP compartments (Welch Two Sample t-test and Wilcoxon rank sum test with continuity correction). e Density of CP DCs in control and progressive MS (Wilcoxon rank sum test). f Density of CP DCs in the different CP compartments (Wilcoxon rank sum test and Welch Two Sample t-test). Col IV: collagen IV; PMS: progressive MS. Scale bars are 100 μm

Fig. 2

T cell density is higher in the CP stroma of progressive MS patients than in the control CP, and they interact with APCs. a Representative images of the control and progressive MS CP immunolabeled with CD3 and collagen IV. White arrowheads point to stromal CD3+ T cells. Scale bars are 100 μm. b Density of CP T cells in the CP of control and progressive MS cases (Welch Two Sample t-test). c Density of CP T cells in the different CP compartments (Wilcoxon rank sum test). d Representative image of a T cell (CD3+, green) in close contact with an APC (MHCII+, red); vessels are visualized with UEA I (white). Maximum projection is accompanied by the orthogonal views. Scale bar is 10 μm. e Absolute density of T cells in close contact with MHCII+ APCs in the CP of control and progressive MS patients, calculated by applying the percentage of T cells that were interacting with APCs to the total density of T cells in their respective samples (Welch Two Sample t-test). f Percentage of T cells interacting with APCs in the CP of control and progressive MS patients, defined as the CP T cells located directly adjacent to MHCII+ cells (Welch Two Sample t-test). g Lack of correlation between interacting T cells and the total CP T cells in each sample (Pearson’s correlation). PMS: progressive MS

Fig. 3

CD8+ T cell density is significantly higher in the CP stroma of progressive MS patients relative to that of controls. a Representative images of the progressive MS and control CP immunolabeled with CD4 (green), CD8 (red) and UEA I (white). White arrows point to CD4+ T cells, while white arrowheads point to CD8+ T cells. b Density of CP CD4+ and CD8+ T cells in the CP of control and progressive MS patients (Wilcoxon rank sum test). Scale bars are 100 μm. c Representative image of a CD4+ T cell (green) and a CD8+ T cell (red) in close contact with an APC (MHCII+, white). Maximum projection is accompanied by the orthogonal views. Scale bar is 10 μm. PMS: progressive MS

Fig. 4

B and plasma cells are virtually absent from the CP. a Examples of CD19+ and CD138+ cells in the CP of progressive MS patients; basement membrane was immunolabeled with collagen IV. White arrowhead in the top panel points to a CD19+ CD138+ cell; in the lower panel, a CD19+ B cell is seen. b Density of CP CD19+ cells in the different CP compartments (Wilcoxon rank sum test with continuity correction). c Density of CP CD138+ cells in the different CP compartments (Wilcoxon rank sum test with continuity correction). PMS: progressive MS. Scale bars are 50 μm

Fig. 5

Granulocyte density is higher in the CP of progressive MS patients relative to the control CP. a Maximum projection image of a progressive MS and a control CP immunolabeled with CD66b (red) and collagen IV (white). White arrowhead points to a granulocyte associated with the epithelium and white arrow points to a stromal granulocyte in the progressive MS case. b Density of CP granulocytes in the CP of control and progressive MS cases (Wilcoxon rank sum test with continuity correction). c Density of CP granulocytes in the different CP compartments (Wilcoxon rank sum test with continuity correction). PMS: progressive MS. Scale bars are 100 μm

Fig. 6

Graphical summary of the immune cell populations in the CP of progressive MS patients and controls. X axis shows the density of each immune cell population in control and progressive MS CP. The area of each bar represents the mean density of the corresponding cell type and disease status. The bar length represents the relative contribution of each disease status to the total cells of a particular subset. Significant alterations in the density of cell populations between progressive MS and control cases are represented with an asterisk. Fold change could not be calculated for B or plasma cells due to the 0 values in the control cases. PMS: progressive MS