Ebook: Handbook of Infection and Alzheimer's Disease

Every four seconds a new case of dementia is diagnosed. Never have people led healthier and longer lives. At a price: this has resulted in a tremendous increase in the number of people with dementia, as dementia mainly affects people when they get very old. But dementia is not a normal part of aging. The total number of people affected increases rapidly and might reach 120 million in 2050. Most of the affected people will be living in Asia, Latin America, or Africa. According to the World Health Organization (WHO), caring for and treating people with dementia currently costs the world more than 600 billion dollars per year.

Alzheimer's disease is the most frequent cause of dementia and has become a major public health concern; in fact the WHO declared dementia a public health priority.

As a previous State Secretary of Research and Education in Switzerland, I am aware of the challenge to governments and health systems, but also to families to care for the growing number of patients suffering from Alzheimer's disease. Finding a cure for this devastating disease is top priority. Many research organizations, thousands of research laboratories and hospitals, various biotech and pharma companies, patient organizations, and foundations have devoted enormous efforts to find therapeutic agents to cure or prevent this terrible disease, with few and disappointing results. Neurodegenerative diseases are so daunting that the most important—and rare—ingredient today is courage to follow new ideas, untrodden paths, and disruptive innovations.

As a trained neurologist, psychiatrist, and neuropathologist, Dr. Judith Miklossy has early on detected that spirochetes (helically shaped bacteria) when they invade brain cells, reproduce the filamentous pathology characteristic of Alzheimer's disease. Dr. Judith Miklossy has defended nearly single-handedly the hypothesis that chronic infection by spirochetes and several other important pathogens could constitute risk factors for Alzheimer's disease.

This is still a debated and largely underfunded alley of research in Alzheimer's disease. But if this hypothesis holds true, early intervention against infection may delay or even prevent the future development of Alzheimer's disease.

As a former State Secretary responsible for science policy, I always wondered how we could foster disruptive, innovative research. Granting agencies, with a well-established (and necessary) peer-review cycle, have trouble funding research for new disruptive ideas. Personalities like Judith, and foundations like the Prevention Alzheimer International Foundation established in Switzerland are the engines of this necessary type of research. Alzheimer's disease is too important and tragic for us to neglect any promising avenue.

Charles Kleiber PhD

State Secretary for Education and Research (1997–2008), Switzerland

Two basic discoveries spurred research into inflammation as a driving force in the pathogenesis of Alzheimer's disease (AD). The first was the identification of activated microglia in association with the lesions. The second was the discovery that rheumatoid arthritics, who regularly consume anti-inflammatory agents, were relatively spared from the disease. These findings led to an exploration of the inflammatory pathways that were involved in AD pathogenesis. A pivotal advance was the discovery that amyloid-β protein (Aβ) activated the complement system. This focused attention on anti-inflammatories as blockers of complement activation. More than 15 epidemiological studies have since showed a sparing effect of non-steroidal anti-inflammatory drugs (NSAIDs) in AD. A consistent finding has been that the longer the NSAIDs were used prior to clinical diagnosis, the greater the sparing effect. The reason has since emerged from studies of biomarkers such as amyloid-β (Aβ) levels in the cerebrospinal fluid and Aβ deposits in brain. They have established that the onset of AD commences at least a decade before cognitive decline permits clinical diagnosis. Such biomarker studies have revealed that a huge window of opportunity exists when application of NSAIDs, other anti-inflammatory agents, or complement activation blockers, could arrest further progress of AD, thus eliminating its manifestation. It can be anticipated that this principle will apply to many other chronic neurodegenerative diseases. Neuroinflammation, discovered in AD more than 30 years ago, has now become a major field of brain research today. Inhibiting it may be the key to successful treatment of many chronic neurological disorders.

Inflammasomes are responsible for the maturation of pro-inflammatory cytokines such as interleukin (IL)-1β, IL-18, and IL-33 and activation of inflammatory cell death, pyroptosis. They assemble in response to cellular infection and stress or to tissue damage, promote inflammatory reactions, and are important in regulating innate immunity particularly by acting as platforms for activation of caspase proteases. They appear to be involved in several pathological processes activated by microbes including Alzheimer's disease (AD). Best characterized in microbial pathogenesis is the nucleotide-binding domain and leucine-rich repeat (NLR)-protein 3 (NLRP3) inflammasome. AD is a neurodegenerative condition in which the neuropathological hallmarks are the deposition of amyloid-β (Aβ)and hyperphosphorylated tau protein coated neurofibrillary tangles. For decades, the role of the innate immune system in the etiology of AD was considered less important, but the recently discovered inflammatory genes by genome-wide association studies driving inflammation in this disease has changed this view. Innate immune inflammatory activity in the AD brain can result from the pathological hallmark protein Aβ as well as from specific bacterial infections that tend to possess weak immunostimulatory responses for peripheral blood myeloid cell recruitment to the brain. The weak immunostimulatory activity is a consequence of their immune evasion strategies and survival. In this review we discuss the possibility that inflammasomes, particularly via the NLR family of proteins NLRP3 are involved in the pathogenesis of AD. In addition, we discuss the plausible contribution of specific bacteria playing a role in influencing the activity of the NLRP3 inflammasome to AD progression.

Alzheimer and a number of other neurodegenerative diseases are characterized by the presence of reactive microglia and reactive astrocytes in association with the lesions. The classic view that microglia exist primarily in either a resting or activated state needs to be broadened in view of recent results. Resting microglia are in constant activity sampling their surround. Activated microglia may be pro-inflammatory, releasing inflammatory cytokines and other inflammatory mediators, or anti-inflammatory, promoting the healing process. There is evidence that microglial phagocytosis is more powerful in the anti-inflammatory state. Activated astrocytes also have pro-inflammatory and anti-inflammatory properties. In the pro-inflammatory state they release inflammatory cytokines. In the anti-inflammatory state they release various growth factors. In AD and other neurodegenerative diseases, both microglia and astrocytes are in a pro-inflammatory state. From a therapeutic point of view it is desirable to find methods of tipping the balance towards an anti-inflammatory state for both types of glia.

Sporadic, late-onset Alzheimer's disease (LOAD) is a progressive neurodegenerative disease that is now the most common and severe form of dementia in the elderly. That dementia is thought to be a direct result of neuronal damage and loss associated with accumulations of abnormal protein deposits in the brain. Great strides have been made in the past 20 years with regard to understanding the pathological entities that arise in the AD brain, both for familial AD (~5% of all cases) and LOAD (~95% of all cases). The neuropathology observed includes: neuritic senile plaques (NSPs), neurofibrillary tangles (NFTs), neuropil threads (NPs), and often deposits of cerebrovascular amyloid. Genetic, biochemical, and immunological analyses have provided a relatively detailed knowledge of these entities, but our understanding of the “trigger” events leading to the biological processes resulting in this pathology and neurodegeneration remains limited. For this reason, the etiology of AD, in particular LOAD, has remained elusive. However, a number of recent and ongoing studies have implicated infection in the etiology and pathogenesis of LOAD. This review focuses specifically on infection with Chlamydophila (Chlamydia) pneumoniae in LOAD and how this infection may function as a “trigger or initiator” in the pathogenesis of this disease.

Alzheimer's disease (AD) is associated with dementia, brain atrophy and the aggregation and accumulation of a cortical amyloid-β peptide (Aβ). Chronic bacterial infections are frequently associated with amyloid deposition. It had been known from a century that the spirochete Treponema pallidum can cause dementia in the atrophic form of general paresis where. It is noteworthy that the pathological hallmarks of this atrophic form are similar to those of AD. Recent observations showed that bacteria, including spirochetes contain amyloidogenic proteins and also that Aβ deposition and tau phosphorylation can be induced in vitro or in vivo following exposure to bacteria or LPS. Bacteria or their poorly degradable debris are powerful inflammatory cytokine inducers, activate complement, affect vascular permeability, generate nitric oxide and free radicals, induce apoptosis and are amyloidogenic. All these processes are involved in the pathogenesis of AD. Old and new observations, reviewed here, indicate that to consider the possibility that bacteria, including several types of spirochetes highly prevalent in the population at large or their persisting debris may initiate cascade of events leading to chronic inflammation and amyloid deposition in AD is important, as appropriate antibacterial and antiinflammatory therapy would be available to prevent dementia.

The cause, or causes, of the vast majority of Alzheimer's disease cases are unknown. A number of contributing factors have been postulated, including infection. It has long been known that the spirochete Treponema pallidum, which is the infective agent for syphilis, can in its late stages cause dementia, chronic inflammation, cortical atrophy and amyloid deposition. Spirochetes of unidentified types and strains have previously been observed in the blood, CSF and brain of 14 AD patients tested and absent in 13 controls. In three of these AD cases spirochetes were grown in a medium selective for Borrelia burgdorferi. In the present study, the phylogenetic analysis of these spirochetes was made. Positive identification of the agent as Borrelia burgdorferi sensu stricto was based on genetic and molecular analyses. Borrelia antigens and genes were co-localized with beta-amyloid deposits in these AD cases. The data indicate that Borrelia burgdorferi may persist in the brain and be associated with amyloid plaques in AD. They suggest that these spirochetes, perhaps in an analogous fashion to Treponema pallidum, may contribute to dementia, cortical atrophy and amyloid deposition. Further in vitro and in vivo studies may bring more insight into the potential role of spirochetes in AD.

Reports that Lyme disease (LD) causes Alzheimer's disease (AD) have appeared in academic journals and online. If the biological agent Borrelia burgdorferi that causes LD also causes AD then areas with highest levels of LD should have significantly higher numbers of deaths due to AD compared to low LD areas. Here we show there is no statistically significant correlation between the incidence of LD and deaths due to AD in the US. Furthermore, the 13 states with the highest deaths due to AD were statistically different (P<0.0001) from those with high LD incidence. Recent work by several other research groups has validated this conclusion.

Alzheimer's disease (AD) is an infectious disease caused by spirochetes,and these spirochetes form biofilms, which attract the innate immune system. The innate immune system first responder, Toll-like receptor 2, generates both NF-κB and TNF- which try to kill the spirochetes in the biofilm, but cannot penetrate the “slime”. NF-κB is also responsible for the generation of amyloid-β (Aβ) which itself is anti-microbial. Aβ cannot penetrate the biofilm either, and its accumulation leads to destruction of the cerebral neurocircuitry. Treatment with penicillin (as in tertiary syphilis, the comparator to AD) is outlined; a biofilm dispersing agent may need to be added to the protocol.

It has long been known that spirochetes form clumps or micro colonies in vitro and in vivo. Cortical spirochetal colonies in syphilitic dementia were considered as reproductive centers for spirochetes. Historic and recent data demonstrate that senile plaques in Alzheimer's disease (AD) are made up by spirochetes. Spirochetes, are able to form biofilm in vitro. Senile plaques are also reported to contain elements of biofilm constituents. We expected that AβPP and Aβ (the main components of senile plaques) also occur in pure spirochetal biofilms, and bacterial DNA (an important component of biofilm) is also present in senile plaques. Histochemical, immunohistochemical, and in situ hybridization techniques and the TUNEL assay were used to answer these questions. The results obtained demonstrate that Aβ and DNA, including spirochete-specific DNA, are key components of both pure spirochetal biofilms and senile plaques in AD and confirm the biofilm nature of senile plaques. These results validate previous observations that AβPP and/or an AβPP-like amyloidogenic protein are an integral part of spirochetes, and indicate that bacterial and host derived Aβ are both constituents of senile plaques. DNA fragmentation in senile plaques further confirms their bacterial nature and provides biochemical evidence for spirochetal cell death. Spirochetes evade host defenses, locate intracellularly, form more resistant atypical forms and notably biofilms, which contribute to sustain chronic infection and inflammation and explain the slowly progressive course of dementia in AD. To consider co-infecting microorganisms is equally important, as multi-species biofilms result in a higher resistance to treatments and a more severe dementia.

The aim of this study was to establish a link between periodontal disease and Alzheimer's disease (AD) with a view to identifying the major periodontal disease bacteria (Treponema denticola, Tannerella forsythia, and Porphyromonas gingivalis) and/or bacterial components in brain tissue from 12h postmortem delay. Our request matched 10 AD cases for tissue from Brains for Dementia Research alongside 10 non-AD age-related controls with similar or greater postmortem interval. We exposed SVGp12, an astrocyte cell line, to culture supernatant containing lipopolysaccharide (LPS) from the putative periodontal bacteria P. gingivalis. The challenged SVGp12 cells and cryosections from AD and control brains were immunolabeled and immunoblotted using a battery of antibodies including the anti-P. gingivalis-specific monoclonal antibody. Immunofluorescence labeling demonstrated the SVGp12 cell line was able to adsorb LPS from culture supernatant on its surface membrane; similar labeling was observed in four out of 10 AD cases. Immunoblotting demonstrated bands corresponding to LPS from P. gingivalis in the SVGp12 cell lysate and in the same four AD brain specimens which were positive when screened by immunofluorescence. All controls remained negative throughout while the same four cases were consistently positive for P. gingivalis LPS (p=0.029). This study confirms that LPS from periodontal bacteria can access the AD brain during life as labeling in the corresponding controls, with equivalent/longer postmortem interval, was absent. Demonstration of a known chronic oral-pathogen-related virulence factor reaching the human brains suggests an inflammatory role in the existing AD pathology.

Periodontal disease is a polymicrobial inflammatory disease that leads to chronic systemic inflammation and direct infiltration of bacteria/bacterial components, which may contribute to the development of Alzheimer's disease. ApoE^−/− mice were orally infected (n=12) with Porphyromonas gingivalis, Treponema denticola, Tannerella forsythia, and Fusobacterium nucleatum as mono-and polymicrobial infections. ApoE^−/− mice were sacrificed following 12 and 24 weeks of chronic infection. Bacterial genomic DNA was isolated from all brain tissues except for the F. nucleatum mono-infected group. Polymerase chain reaction was performed using universal 16s rDNA primers and species-specific primer sets for each organism to determine whether the infecting pathogens accessed the brain. Sequencing amplification products confirmed the invasion of bacteria into the brain during infection. The innate immune responses were detected using antibodies against complement activation products of C3 convertase stage and the membrane attack complex. Molecular methods demonstrated that 6 out of 12 ApoE^−/− mice brains contained P. gingivalis genomic DNA at 12 weeks (p=0.006), and 9 out of 12 at 24 weeks of infection (p=0.0001). Microglia in both infected and control groups demonstrated strong intracellular labeling with C3 and C9, due to on-going biosynthesis. The pyramidal neurons of the hippocampus in 4 out of 12 infected mice brains demonstrated characteristic opsonization with C3 activation fragments (p=0.032). These results show that the oral pathogen P. gingivalis was able to access the ApoE^−/− mice brain and thereby contributed to complement activation with bystander neuronal injury.

This review shows how our microbiome influences health and ultimately how well we age. Evidence linking oral bacteria to Alzheimer's disease (AD) is discussed in the context of aging, drawing together data from epidemiological, experimental, genetic and environmental studies. Immunosenescence results in increased bacterial load as cell-mediated and humoral immune responses wane, with the innate immune system contributing to a rise in circulating proinflammatory cytokines such as TNF and IL1β. Aging may favor the proliferation of anaerobes in the mouth eliciting a robust TNF response from the oral epithelium. Maintaining the integrity of the blood-brain-barrier (BBB) against a backdrop of increasing bacterial load is important; prolonged exposure to high levels of TNF compromises its integrity. Sensitive techniques now detect the “asymptomatic” presence of bacteria in areas previously thought as sterile, providing new insights into the wider distribution of components of the microbiome. These “immune-tolerated” bacteria may slowly multiply elsewhere until they elicit a chronic inflammatory response; some being considered causal in instances of atherosclerosis and back pain. Inflammatory processes, long associated with AD, have recently been further elucidated, in particular revealing the role of the inflammasomes. We propose for a subset of AD patients, aging favors the overgrowth of oral anaerobes, established earlier in life, provoking a pro-inflammatory innate response that weakens the BBB allowing bacteria to spread and quietly influence the pathogenesis of AD. Finally, we suggest that human polymorphisms, considered alongside components of the microbiome, may provide new avenues of research for the prevention and treatment of disease.

Background: The possibility of an infectious etiology for Alzheimer's disease (AD) has been repeatedly postulated over the past three decades, with the roles of both viruses and bacteria having been investigated. Chlamydophila (formerly Chlamydia) pneumoniae (Cpn) and spirochetal bacteria have been two of the most frequently implicated bacterial groups in AD pathogenesis.

Objective: A meta-analysis was performed where data were combined from 25 studies examining the association between AD and spirochetal bacteria or Cpn.

Methods: Comprehensive search of several electronic databases. Data were extracted from published studies and a random-effects model was used to analyze the data.

Results: A statistically significant association between AD and detectable evidence of infection of either bacterial group was demonstrated. Over a ten-fold increased occurrence of AD was noted when there is detectable evidence of spirochetal infection (OR: 10.61; 95% CI: 3.38–33.29), with a more conservative risk estimate demonstrating over a four-fold increased occurrence of AD (OR 4.45; 95% CI: 2.33–8.52). Over a five-fold increased occurrence of AD was noted with Cpn infection (OR 5.66; 95% CI: 1.83–17.51).

Discussion: There appears to be a strongly positive association between bacterial infection and AD.

Alzheimer's disease (AD) affects approximately 5.3 million people in the U.S. and this number will increase as the population ages and the life-span increases. Therefore, of paramount importance is identifying mechanisms and factors that affect the risk of developing AD. The etiology and pathogenic mechanisms for AD have not been defined, although inflammation within the brain is thought to play a significant role. Consistent with this hypothesis, studies suggest that peripheral inflammations, dysbiotic conditions, and infections contribute to the inflammatory state of the brain and may constitute risks for AD. Recently, several peripheral conditions with an inflammatory basis such as diabetes and obesity have been recognized as risks for AD. Periodontitis is a prevalent, chronic peripheral polymicrobial disease associated with gram negative, anaerobic bacteria, which exhibits significant localized and systemic inflammatory effects. This review will present evidence suggesting that periodontal disease may also be a risk factor for AD and possible mechanistic links between periodontitis related inflammation and AD. It will review the pathogenesis of periodontitis and the mechanisms by which periodontal infections may affect the onset and progression of AD. Periodontitis is a treatable condition and may be a readily modifiable risk factor for AD. Therefore, further studies including intervention trials are warranted.

The mechanisms of disease processes resulting in dementia of which, Alzheimer's disease (AD) is a common example, remain elusive. To this end, a number of theories as plausible explanations have been suggested. Of these, the microbial, peripheral infection theory of Hunter and Miller (1900s) and Naguchi and Moore (1913) is the earliest proposal to explain possible causation of AD. Periodontal disease is a polymicrobial inflammatory disease reported to associate with AD via periodontal bacteria/bacteraemia, systemic inflammation, blood-brain barrier erosion, intra-cerebral inflammation and tissue injury, this chapter describes the original finding of four out of 10 confirmed AD brains with incidental infection of Porphyromonas gingivalis [P. gingivalis] outer membrane component lipopolysaccharide. A follow-on study examined the possibility of P. gingivalis translocation from the gingivae to the brain in the orally infected (n=12), apoliporprotein E knockout (ApoE^−/−) mouse model at 12 and 24 weeks of monoinfections. Sensitive bacterial molecular speciation techniques confirmed the invasion of P. gingivalis into the brain at 12 weeks (p=0.006), and at 24 weeks of infection (p=0.0001). Immunolabeling using antibodies against complement proteins demonstrated the innate immune system activation via C3 fragmentation and its subsequent opsonisation onto vulnerable pyramidal neurons (p=0.032) in the hippocampus as ongoing bystander injury. These studies confirm the initiation of an infection mediated inflammasome assembly with implications for remote body organ inflammatory pathologies from periodontitis to dementia.

The last 8 or so years have seen a large increase in the number of studies supporting the concept of a major role for herpes simplex virus type 1 (HSV1) in Alzheimer's disease (AD). The main advances have been made through studies in humans and in mice, investigating the likelihood of reactivation of the latent virus in brain. Others have aimed to explain the mechanisms in cells whereby the increase in amyloid-beta (Aβ) production on HSV1 infection of cells and mouse brains occurs, and the reason that infected cells make this increase. The possibility that other herpesviruses are involved in the development of AD has been explored, and human herpesvirus type 6, Epstein-Barr virus, and cytomegalovirus, in particular, have been implicated. Epidemiological studies have further supported the role specifically of HSV1 and its reactivation in the disease. Antiviral studies have continued, comparing those acting by different mechanisms, such as restricting viral replication, or blocking viral entry into cells, to treat HSV1-infected cell cultures, and then examining the extent to which the virus-induced increases in Aβ and AD-like tau are reduced. All the studies support the usage of antiviral treatment to slow or halt the progression of AD.

Herpes simplex virus type 1 (HSV-1) is a neurotropic virus able to establish a persistent latent infection in the host. Herpes simplex encephalitis (HSE) is associated with a high mortality rate and significant neurological, neuropsychological, and neurobehavioral sequelae, which afflict patients for life. Currently, it is unclear whether asymptomatic recurrent reactivations of HSV-1 occur in the central nervous systems in infected people, and if these events could lead to a progressive deterioration of neuronal function. In this context, HSV-1 constitutes an important candidate to be included among the risk factors for the development of Alzheimer's disease. Our group have demonstrated that HSV-1 triggers neurodegenerative events in in vitro and in vivo induced neuronal infection, evidenced by increase in tau hyperphosphorylation and caspase-3 dependent cleavage of tau protein, resembling what occurs in neurodegenerative diseases. In addition, in an in vivo model, a reactivation episode during asymptomatic latency of HSV-1 infection in mice was accompanied by upregulation of neuroinflammatory markers (toll-like receptor-4, interferon /β, and p-IRF3). Besides, previous reports have shown that HSV-1 inhibits apoptosis during early infection, but is pro-apoptotic during productive infection. Taking in consideration that the stress sensors AMPK and Sirt1 are involved in neuronal survival and neuroprotection, we hypothesized that HSV-1 could activate the AMPK/Sirt1 axis as a strategy to establish latency through inhibition of apoptosis and restoration of the energy status. Thus, we demonstrated that HSV-1 modulates the AMPK/Sirt1 axis differentially during infection, interfering with pro-apoptotic signaling and regulating mitochondrial biogenesis, pivotal processes in the lifetime of neurons in the central nervous system. In conclusion, our findings support the idea that HSV-1 could contribute to induce neurodegenerative processes in age-associated pathologies such as Alzheimer's disease.

Amyloid-β (Aβ) peptides generated by the amyloidogenic pathway of amyloid-β protein precursor processing contribute significantly to neurodegeneration characteristic of Alzheimer's disease (AD). The involvement of Aβ peptides in the etiology of AD remains a subject of debate. Data published in the last 6 years by three different groups have added a new twist by revealing that Aβ peptides could act as antimicrobial peptides (AMP) in in vitro assays against some common and clinically relevant microorganisms,inhibit replication of seasonal and pandemic strains of influenza A and HSV-1 virus. These observations are of significance with respect to the notion that pathogens may be important contributors to the development of AD, particularly in the case of herpes simplex virus (HSV) infection, which often resides in the same cerebral sites where AD arises. Here, we review the data that support the interpretation that Aβ peptides behave as AMP, with an emphasis on studies concerning HSV-1 and a putative molecular mechanism that suggests that interactions between Aβ peptides and the HSV-1 fusogenic protein gB lead to impairment of HSV-1 infectivity by preventing the virus from fusing with the plasma membrane. A number of avenues for future research are suggested.