Photodynamic opening of the blood-brain barrier affects meningeal lymphatics and the brain's drainage in healthy male mice

Abstract

Here, we present the new vascular effects of photodynamic therapy (PDT) with 5-aminolevulinic acid (5-ALA). PDT with 5-ALA induces a leakage of both the meningeal and cerebral blood vessels. The extravasation of photo-excited 5-ALA from the leaky blood vessels into the meninges causes photo-damage of the meningeal lymphatics (MLVs) leading to a dramatic reducing the MLV network and brain's drainage. The PDT-induced impairment of lymphatic regulation of brain's drainage can lead to excessive accumulation of fluids in brain tissues, which is important to consider in the PDT therapy for brain diseases as s possible side effect of PDT with 5-ALA.

Fig. 1.

The PDT effects on FITCD distribution in the brain and its accumulation in dcLNs: (a and b) Representative images of the bottom aspect of the brain (a) and dcLNs (b) before PDT in the control groups, including mice treated with 5-ALA (i.c.m.), 5-ALA (i.v.) or laser alone and 7 days after PDT in the 5-ALA (i.c.m.) + Laser and the 5-ALA (i.v.) + Laser groups; (с and d) Quantitative analysis of the signal intensity from FITCD in the brain (c) and in dcLNs (d) in the tested groups, n = 8 in each group, * - p < 0.05, ** - p < 0.01, *** - p < 0.001, the ANOVA test with post hoc Duncan test.

Fig. 2.

The PDT effects on the dorsal MLV network and the permeability of the meningeal blood vessels: (a) Representative images of LYVE-1 + vessels (green) covering the main venous sinuses (SSS and TS) in the control and PDT groups. The blood vessels are marked with CD 31 (red) and filled with Evans Blue (blue), DAPI (violet); (b) the schematic illustration of ROI for z confocal analysis of the dorsal MLVs; (c) Quantitative analysis of LYVE-1 coverage expressed in % of ROI, n = 8 in each group, * - p < 0.05, *** - p < 0.001, the ANOVA test with post hoc Duncan test.

Fig. 3.

The PDT-mediated OBBB: (a and b) Representative images (2D on the left and 3D on the right) of in vivo multiphoton analysis of the cortex before PDT (a) with the intact cerebral vessels and 1 h after PDT with 5-ALA (i.v.) when Evans Blue (red) leakage was observed as red clouds around the cerebral vessels; (c and d) Representative images of ex vivo confocal analysis of the cortex before PDT (c) and 1 h after PDT with 5-ALA (i.v.). The extravasation of Evans Blue (green) was observed as green clouds around the cerebral vessels marked with CD 31 (red), astocytes are marked with GFAP (blue).