Fig. 6: Disrupted LN architecture in the presence of particulate matter.
a, Representative confocal images of whole human LN sections from donors of the indicated ages, stained for CD20 to show B cell follicles (green) and with brightfield imaging overlays (white) to show black particulate matter. Representative images were taken from 3−7 donors (LLNs) and 2–8 donors (MLNs) per age group (≤39, 40–64 and ≥65 years). Scale bars, 1,000 µm. b, B cell circularity measured using an open-source code (see Methods) to quantify B cell follicle integrity in MLNs (n = 14) and LLNs (n = 17). The graphs show the circularity measurement above a threshold (>0.5 µm) for each whole LN section obtained from organ donors of the indicated ages. Linear regression was performed with Pearson’s correlation (two tailed). P values and Pearson’s R values are provided (** for p<0.01). The data are presented as means ± 95% CIs. c, Representative confocal images of human LLNs (from donors of the indicated ages) stained for podoplanin to show the lymphatics (red; middle). Brightfield imaging overlays (white; left) show the black particulate matter. In the right column, they are pseudocolored green to show the overlay with podoplanin (the particulate matter and lymphatics were isosurfaced using Imaris software). Representative images were taken from 5–12 donors per age group (≤65 and ≥65 years). Scale bars, 1,000 µm. d, Left, graph showing the percentage of the whole LN section covered by lymphatics. Right, graph showing the area covered by lymphatics close to the particulate area (<3 µm), as determined using Imaris software. Statistical significance was determined by one-way ANOVA with Tukey’s post-hoc test. Statistically significant differences were found for the following comparisons of podoplanin+ area (right graph): ≤39 versus 40–64 years (P = 0.0257, *) and ≤39 versus ≥65 years (P = 0.0279, *). The data are presented as means ± s.d. and are from 17 donors.
