Primary Cilia Structure Is Prolonged in Enteric Neurons of 5xFAD Alzheimer’s Disease Model Mice

Abstract

Neurodegenerative diseases such as Alzheimer’s disease (AD) have long been acknowledged as mere disorders of the central nervous system (CNS). However, in recent years the gut with its autonomous nervous system and the multitude of microbial commensals has come into focus. Changes in gut properties have been described in patients and animal disease models such as altered enzyme secretion or architecture of the enteric nervous system. The underlying cellular mechanisms have so far only been poorly investigated. An important organelle for integrating potentially toxic signals such as the AD characteristic A-beta peptide is the primary cilium. This microtubule-based signaling organelle regulates numerous cellular processes. Even though the role of primary cilia in a variety of developmental and disease processes has recently been recognized, the contribution of defective ciliary signaling to neurodegenerative diseases such as AD, however, has not been investigated in detail so far. The AD mouse model 5xFAD was used to analyze possible changes in gut functionality by organ bath measurement of peristalsis movement. Subsequently, we cultured primary enteric neurons from mutant mice and wild type littermate controls and assessed for cellular pathomechanisms. Neurite mass was quantified within transwell culturing experiments. Using a combination of different markers for the primary cilium, cilia number and length were determined using fluorescence microscopy. 5xFAD mice showed altered gut anatomy, motility, and neurite mass of enteric neurons. Moreover, primary cilia could be demonstrated on the surface of enteric neurons and exhibited an elongated phenotype in 5xFAD mice. In parallel, we observed reduced β-Catenin expression, a key signaling molecule that regulates Wnt signaling, which is regulated in part via ciliary associated mechanisms. Both results could be recapitulated via in vitro treatments of enteric neurons from wild type mice with A-beta. So far, only a few reports on the probable role of primary cilia in AD can be found. Here, we reveal for the first time an architectural altered phenotype of primary cilia in the enteric nervous system of AD model mice, elicited potentially by neurotoxic A-beta. Potential changes on the sub-organelle level—also in CNS-derived neurons—require further investigations.

5xFAD mice do not have myenteric amyloidosis, dysregulation of neuromuscular transmission or gastrointestinal dysmotility

“…More importantly, the authors have analyzed the expression of the most common AD‐linked genes in different regions of the GI tract, but overall they did not find significant changes in these gene expressions in 5xFAD mice compared with WT mice, even until at 40 weeks of age 27 . In their studies, even the expression of APP and PSEN1/2 genes in entire GI tract were very comparable between 5xFAD vs WT mice, 27 these data against that overexpressions of APP/PSEN gene have been employed in GI from 5xFAD mice, but the data are strongly supporting our observations that 5xFAD mice do not have amyloidosis in colonic ENS, since 5xFAD mice should be with the overexpression of three APP and two PSEN1 genes.…”

“…mice cannot be concluded without the support of amyloidosis in ENS. In previous studies in 5xFAD mice, 22,27 the authors did not provide any direct evidence showing the occurrence of amyloidosis in GI from 5xFAD mice. More importantly, the authors have analyzed the expression of the most common AD-linked genes in different regions of the GI tract, but overall they did not find F I G U R E 6 Detection of NO release from colonic myenteric ganglia in WT and 5xFAD male and female mice using continuous amperometry.…”

“…In AD patients, Aβ accumulation in the gut wall and stool have been reported, 14–18 which strongly supports the contribution of amyloidosis in GI dysmotility of AD patients. Some mouse models of AD also revealed amyloidosis in the GI wall and moderation of GI dysmotility, alterations of microbiome composition, disturbances in gut permeability, changes GI structure and systemic inflammation 15,20–23,27 . However, Aβ accumulation associated with enteric neuronal dysfunction, specifically myenteric neurons in GI dysmotility, has not been studied mechanistically, even though Aβ accumulation was directly detected by positive immune staining in the myenteric ganglia in AD mouse models of AβPP/PS1 (APPswe, PSNE1dE9) 15 and TgCRND8, 23 .…”

Injection of amyloid-β to lateral ventricle induces gut microbiota dysbiosis in association with inhibition of cholinergic anti-inflammatory pathways in Alzheimer’s disease

“…Furthermore, Aβ 1-42 i.c.v can induce changes in intestinal structure, including colon length, intestinal mucosal barrier integrity, and enteric neuron number in mice. Previous studies have shown similar changes in intestinal structure and function in transgenic AD model mice [8,37]. For example, Puig et al reported that occludin, an epithelial tight junction protein, were overall decreased in small intestine of APP/PS1 mice [8].…”

“…For example, Puig et al reported that occludin, an epithelial tight junction protein, were overall decreased in small intestine of APP/PS1 mice [8]. In addition to the changes of gut anatomy and motility, there were also elongated phenotypes of primary cilia in 5xFAD mice [37].…”