# PDE4B deficiency aids macrophage differentiation and contributes to Cryptococcus neoformans brain infection

## TL;DR

This study shows that PDE4B regulates macrophage behavior during fungal brain infections, suggesting it could be a new treatment target for cryptococcal meningitis.

## Contribution

The study identifies PDE4B as a novel regulator of macrophage-mediated fungal dissemination in cryptococcal brain infection.

## Key findings

- PDE4B inhibition increases macrophage expression of Arg1, CXCR4, and CCR7, promoting fungal crossing of the blood-brain barrier.
- PDE4B knockout or inhibition worsens brain fungal burden, which is partially reversed by macrophage depletion.
- PDE4B activation reduces fungal burden in the brain even after infection onset.

## Abstract

Cryptococcal meningitis is a fatal complication. Macrophages have been proposed to function as candidate “Trojan horse” cells, transferring Cryptococcus neoformans (C. neoformans) into the brain. The mechanisms of Trojan horses in cryptococcal meningitis are largely elusive. In this study, we performed scRNA-Seq on immune cells infiltrating the brain in a murine model of cryptococcal meningitis. Bioinformatics analysis revealed that phosphodiesterase 4B (PDE4B) is a candidate regulator associated with C. neoformans infected-macrophage. C. neoformans increases the total level of PDE4B in macrophages. However, virulent strains with increased production of melanin paradoxically decreased PDE4B expression in macrophages, implying that PDE4B in macrophages may be negatively associated with C. neoformans invasion. PDE4B inhibition increased Arg1, CXCR4 and CCR7 expression in macrophages, a process regulated by the cAMP/PKA signaling pathway. As expected, PDE4B inhibitors promote the ability of C. neoformans infected-macrophages to cross the blood–brain barrier (BBB) in vitro. Similarly, PDE4B inhibitors or PDE4B knockout increase the fungal burden in the brain, which is, at least partially, rescued by macrophage depletion, and adoptive transfer experiments further support macrophage-mediated fungal delivery to the brain. In contrast, PDE4B activation reduces fungal burden in the brain, including when administered after infection onset. Overall, this study revealed that PDE4B functions as an important regulator of macrophage functional programming during infection and supports a macrophage-mediated dissemination mechanism contributing to brain invasion, and is a potential therapeutic target for cryptococcal meningitis.

Cryptococcus neoformans (C. neoformans), a facultative intracellular fungus, is the pathogen of destructive cryptococcal meningoencephalitis. Macrophages have been proposed to function as candidate “Trojan horse” cells, transferring C. neoformans across the blood‒brain barrier (BBB), although definitive in vivo evidence remains limited. Using unbiased scRNA-seq profiling of brain immune cells from C. neoformans-infected mice, we revealed that PDE4B is a candidate regulator associated with macrophage differentiation and migratory responses upon C. neoformans brain infection. In vitro, PDE4B inhibitors or PDE4B knockout promote the crossing of the BBB by C. neoformans-infected macrophages. In vivo, PDE4B inhibitors or PDE4B knockout aggravate C. neoformans brain infection, which is at least partially reversed by macrophage depletion. And adoptive transfer experiments further support macrophage-mediated fungal delivery to the brain. In contrast, PDE4B activation reduces fungal burden in the brain, including when administered after infection onset. PDE4B therefore emerges as a potential host-directed therapeutic target for the prevention and treatment of cryptococcal meningoencephalitis. Considering that PDE4B inhibitors have been widely used in clinical practice, our findings highlight the need for caution in their use in patients at risk for cryptococcal infection.