Extended Data Fig. 4: PICALM expression in human postmortem brains.

(a) Human post-mortem brain sections from an asymptomatic patient were stained with antibodies against PICALM (green) and IBA1 (magenta). White arrows indicate PICALM expressed in MG, and yellow arrows indicate PICALM expressed in a blood vessel. Line-scan (kymography) analysis (right) demonstrates the overlapping expression. The white line in IBA1 composite image shows the location of the line scan. Scale bar, 50 µm. (b) Reduced PICALM expression in the grey matter of AD patients’ postmortem brains (n = 20) compared to controls (n = 19). Two-sided unpaired Student’s t-test was used. (c) Comparison of brain MG state-specific marker gene expression between different human brain MG states. MG states were defined as described in the snRNA-seq study¹⁹. Z-scored expression values of MG state-specific markers and PICALM are shown. HOMEO, homeostatic MG including cell clusters 1,2,3,5 and 8; DAM, disease-associated MG including clusters 0 and 7; LDAM, LD-accumulative MG including clusters 4 and 6. Note lower PICALM expression in DAM and LDAM cell clusters. (d) Representative immunofluorescence staining images of post-mortem brain sections from unaffected control (CERAD 0) and AD (CERAD 3) patients. Green arrows in single-channel images indicate IBA1+ microglia which are also positive for PICALM. CERAD = Consortium to Establish a Registry for AD. (e) Representative immunofluorescence staining of PICALM in individual microglia from Braak stages 1–5 and CERAD scores 0–3, quantified in e-f. Scale bar 20 µm. (f) Quantification of PICALM expression in MG of AD and non-AD controls. AD, n = 15 with the diagnosis of “definitely AD” with a CERAD score of 3; the rest include control (n = 16), probable AD and possible AD (n = 9) (Supplementary Table 11). The grey background dots represent individual MG. ns, not significant in the two-sided unpaired Student’s t-test. (g) Quantification of PICALM expression in MG of AD and non-AD controls stratified by Braak Stage (0-5) (left) and by CERAD score (0 = control; 1-2, probable or possible AD; and 3, definitely AD) (right). Not significant in one-way ANOVA. (h) PICALM protein abundance in postmortem brain prefrontal cortex between AD, asymptomatic AD (AsymAD), and non-AD individuals. Data are from a published dataset²⁷. One-way ANOVA; Dunnett’s correction. In all comparisons. * P < 0.05, ** P < 0.01, ***, P < 0.001; ns=not significant; mean ± s.e.m.