Gram-negative bacterial molecules associate with Alzheimer disease pathology

Abstract

Objective:We determined whether Gram-negative bacterial molecules are associated with Alzheimer disease (AD) neuropathology given that previous studies demonstrate Gram-negative Escherichia coli bacteria can form extracellular amyloid and Gram-negative bacteria have been reported as the predominant bacteria found in normal human brains.Methods:Brain samples from gray and white matter were studied from patients with AD (n = 24) and age-matched controls (n = 18). Lipopolysaccharide (LPS) and E coli K99 pili protein were evaluated by Western blots and immunocytochemistry. Human brain samples were assessed for E coli DNA followed by DNA sequencing.Results:LPS and E coli K99 were detected immunocytochemically in brain parenchyma and vessels in all AD and control brains. K99 levels measured using Western blots were greater in AD compared to control brains (p < 0.01) and K99 was localized to neuron-like cells in AD but not control brains. LPS levels were also greater in AD compared to control brain. LPS colocalized with Aβ1-40/42 in amyloid plaques and with Aβ1-40/42 around vessels in AD brains. DNA sequencing confirmed E coli DNA in human control and AD brains.Conclusions:E coli K99 and LPS levels were greater in AD compared to control brains. LPS colocalized with Aβ1-40/42 in amyloid plaques and around vessels in AD brain. The data show that Gram-negative bacterial molecules are associated with AD neuropathology. They are consistent with our LPS-ischemia-hypoxia rat model that produces myelin aggregates that colocalize with Aβ and resemble amyloid-like plaques.

A systemic view of Alzheimer disease — insights from amyloid-β metabolism beyond the brain

“…Gram-negative bacterial species (such as Escherichia coli K99) are the predominant sources of bacteria-derived factors in normal human brains, and levels of these molecules are increased in AD brains, along with levels of the bacterial cell wall component lipopolysaccharide 128 . Lipopolysaccharide colocalizes with Aβ in plaques in AD brains 128 , suggesting that Gram-negative bacteria are associated with AD pathogenesis. Probiotic supplementation is associated with improved cognition in patients with AD 129 , which further supports a role for the gut microbiota in AD development.…”

Gastrointestinal (GI) Tract Microbes and Microbial Neurotoxins in the Human Central Nervous System (CNS) in Alzheimer’s Disease (AD)

“…Of further related interest are the recent observations: (i) that the increased abundance of gram-negative GI tract bacteria such as B. fragilis in AD patients appears to result in increased generation and translocation of LPS and other Bacteroides-derived neurotoxins from the GI tract into the systemic circulation, which in turn may contribute to AD neuropathology through the release of pro-inflammatory cytokines, systemic inflammation, an increase in GI-tract or BBB permeability or other AD-relevant pathogenic mechanisms [9,[29][30][31][32], and (ii) that an increase in Bacteroidetes in the GI tract is also associated with Parkinson's disease (PD; [33]) and with sporadic AD hippocampus and neocortex, two anatomical regions targeted by the AD process [6,[34][35][36]. Importantly, while only the inflammatory potential of LPS towards primary human neuronal-glial (HNG) co-cultures have been studied and quantified by the induction of the pro-inflammatory NFkB p50/p65 complex, Bacteroidetes species are capable of secreting an unusually complex array of highly lethal neurotoxins including amyloids, sncRNAs and endotoxins which, when released from the confines of the healthy GI tract, are systemically pathogenic and can be highly detrimental to the homeostatic function of human CNS neurons [15].…”

“…As fore-mentioned, microbes such as B. fragilis and Escherichia coli (E. coli), abundant anaerobic Gram-negative bacilli of the human GI-tract microbiome, appear to direct critical regulatory roles of pathogenicity through the stress-induced secretion of a complex mixture of bacterial amyloids, endotoxins and exotoxins, 'microRNA-like' sncRNAs and LPS. Recently work from several independent groups has further described the presence of intact bacteria, bacterial-derived nucleic acid sequences and/or bacterial-derived neurotoxins such as a highly pro-inflammatory LPS that is associated with neuronal parenchyma and in particular the neuronal nuclei of anatomical regions of the ADaffected brain exhibiting characteristic neuropathology [1,9,28,29,[34][35][36][37][38][39]. Interestingly, the close association of bacterial LPS with neuronal nuclei may prevent the efficient export of messenger RNA (mRNA) from a highly active neuronal genome resulting in the down-regulation of gene expression in AD as is widely observed [6,15].…”

“…It has been very recently shown (i) that both LPS and BF-LPS are abundant in anatomical regions of the human brain's limbic system in AD, including the hippocampus and neocortex that exhibit focused neuropathology and an intense inflammatory response as is characteristic of the AD process [6,28,29]; and (ii) that BF-LPS is an extremely potent inducer of pro-inflammatory gene signaling pathways, as quantified by the evolution of the pro-inflammatory transcription factor NF-kB p50/p65 complex HNG cells in primary co-culture [51]. To further cite several highly relevant research investigations: (i) using immunocytochemistry E. coli K99 pili protein and E. coli LPS levels were found to be significantly greater in AD compared to control brains, finding that in AD LPS co-localized with Aβ1-40/42-positive amyloid plaques surrounding cerebral vessels [29]; (ii) using an immunocytochemical approach Zhao et al have discovered LPS in very short post-mortem interval (PMI) AD hippocampus (1-3 h PMI) to levels up to 36-fold over age-matched controls [34][35][36]; (iii) co-incubation of Aβ peptide with LPS potentiates amyloidogenesis and fibrillogenesis (amyloid fiber formation; [34,52]); (iv) systemic injection of LPS in wild-type and transgenic AD mice results in greater amyloid deposition and tau pathology [53]; and (v) a selective enrichment of LPS specifically associated with the neuronal nuclear membrane in AD was found to be correlated to a down-regulation in the output of transcription products from neuronal nuclei ( [34]; manuscript under review). This suggests that GI-tract microbiome-derived LPS may be an important initiator and/or significant contributor to failure in adequate gene expression the AD CNS including those genes involved in the modulation of inflammatory signaling (Figure 1; manuscript submitted).…”

“…Related to the points above, overall the GI tract microbiome of AD patients was found to exhibit highly selective abundance and compositional differences in genus and species from control age-and sex-matched individuals and exhibited phylum-and genus-wide differences in bacterial abundance including a significantly increased abundance of Bacteroidetes and decreased abundance of Firmicutes and Bifidobacterium in the AD-affected brain [9,[14][15][16][17][23][24][25][26]. Very recently, at least 2 independent publications have shown an enrichment of LPS within the neocortex or hippocampus of ADaffected brain [6,[27][28][29].…”