The choroid plexus links innate immunity to CSF dysregulation in hydrocephalus

Highlights

• Integrated multi-omics study of infectious and hemorrhagic hydrocephalus

• Blood or bacteria in CSF elicit highly similar ChP immune and secretory responses

• Crosstalk between ChP immune and epithelial cells drives pathologic CSF secretion

• Immunomodulators treat PIH/PHH by antagonizing a SPAK-regulated ChP transportome

Summary

The choroid plexus (ChP) is the blood-cerebrospinal fluid (CSF) barrier and the primary source of CSF. Acquired hydrocephalus, caused by brain infection or hemorrhage, lacks drug treatments due to obscure pathobiology. Our integrated, multi-omic investigation of post-infectious hydrocephalus (PIH) and post-hemorrhagic hydrocephalus (PHH) models revealed that lipopolysaccharide and blood breakdown products trigger highly similar TLR4-dependent immune responses at the ChP-CSF interface. The resulting CSF “cytokine storm”, elicited from peripherally derived and border-associated ChP macrophages, causes increased CSF production from ChP epithelial cells via phospho-activation of the TNF-receptor-associated kinase SPAK, which serves as a regulatory scaffold of a multi-ion transporter protein complex. Genetic or pharmacological immunomodulation prevents PIH and PHH by antagonizing SPAK-dependent CSF hypersecretion. These results reveal the ChP as a dynamic, cellularly heterogeneous tissue with highly regulated immune-secretory capacity, expand our understanding of ChP immune-epithelial cell cross talk, and reframe PIH and PHH as related neuroimmune disorders vulnerable to small molecule pharmacotherapy.

Introduction

The choroid plexus (ChP) projects into each of the four cerebrospinal fluid (CSF)-filled ventricles and comprises an epithelial cell sheet populated by immune and mesenchymal cell types.^5,6 The ChP is the body’s secretory epithelium par excellence, producing a half-liter of CSF per day via the concerted action of its multiple ion and water transport molecules.⁷ The ChP is also the blood-CSF barrier, gating circulating immune cell entry into the CSF in the setting of infection or tissue damage.^8,9,10,11,12 The ChP’s location at the nexus of the CSF, brain parenchyma, and systemic circulation allows it to sense perturbations and transduce danger signals into homeostatic responses. However, the cellular and molecular mechanisms that coordinate the immune and secretory functions of the ChP are poorly understood.

Hydrocephalus is characterized by the expansion of the cerebral ventricles (ventriculomegaly). Post-infectious hydrocephalus (PIH) and post-hemorrhagic hydrocephalus (PHH), the most common forms of hydrocephalus, are treated by neurosurgical CSF diversion with or without ChP cauterization to decrease CSF production.^13,14,15 These life-saving operations have significant long-term morbidity and failure rates, and are unavailable in impoverished areas.^14,16,17 Furthermore, ChP cauterization may also disrupt normal ChP functions important for brain development and immune function.¹⁸ Drug treatments for hydrocephalus remain unavailable.¹⁸ An improved understanding of ChP biology could identify therapeutic targets for hydrocephalus and other CSF disorders and neuroimmune diseases.^{8,12,18,19,20,21}

PIH and PHH are commonly attributed to intraventricular CSF accumulation secondary to decreased CSF reabsorption due to obstruction of intraventricular CSF flow and/or blockage of the extraventricular arachnoid granulations.²² In contrast, the role of ChP’s immune-secretory functions to the pathogenesis of hydrocephalus remains under-investigated. Interestingly, human PIH and PHH exhibit similar CSF immune cell and cytokine profiles,^23,24 and CSF neutrophilic pleocytosis can predict the development of PHH.²⁵ These observations suggest an important, yet uncharacterized, role of ChP and CSF inflammation in the pathogenesis of acquired hydrocephalus.

Systemic epithelia respond to pro-inflammatory stimuli by increasing rates of fluid transport²⁶ to clear organisms or tissue debris.^27,28,29 However, inappropriately triggered or maladaptively sustained epithelial inflammation can dysregulate ion transport homeostasis. Examples include chemical, autoimmune, and infectious forms of pleuritis, colitis, and pancreatitis.^29,30,31,32 Intraventricular hemorrhage (IVH) is known to cause inflammation-induced ChP CSF hypersecretion in PHH models.^{2,33,34,35,36} Nonetheless, the mechanism(s) by which the ChP contributes to PHH and PIH remain obscure, hindering the development of non-surgical treatment strategies.

Lipopolysaccharide (LPS)-expressing bacteria commonly cause PIH.^37,38,39 LPS is the canonical pathogen-associated molecular pattern (PAMP) for Toll-like receptor-4 (TLR4).^40,41 PHH-derived hemoglobin, and possibly other blood products, are TLR4-stimulating damage-associated molecular patterns (DAMPs).^42,43 We hypothesized that PIH and PHH have convergent pathophysiology that dysregulates ChP immune-secretory function. To test this, we created rat models of PIH and PHH and conducted a multi-omics investigation of these animals to dissect the physiologic, cellular, and molecular pathology of PIH and PHH. Our results suggest that PIH and PHH are related neuroinflammatory disorders amenable to systemic immunomodulation.