Influence of the Gut Microbiota on the Pathogenesis of Alzheimer’s Disease: A Literature Review

The composition of the microbiota is established at the beginning of life, and in healthy individuals, it is relatively similar throughout most of life. With age and the emergence of chronic diseases, toxins, and medications, this composition may be disturbed, and as a result, the older patients are in a chronic low-grade state of inflammation [29], which will be described further.

It is currently estimated that there may be over a thousand species, or in other words, seven thousand strains of bacteria in the human intestines [8,34]. A significant part of the human microbiota, that is up to eighty percent, consists of the phyla Bacteroidetes and Firmicutes. Other phyla worth noting are Actinobacteria, Fusobacteria, and Verrucomicrobia. Typically pathogenic bacteria, such as Campylobacter jejuni, Salmonella enterica, Vibrio cholerae, Escherichia coli, or Bacteroides fragilis, are usually around 0.1% [33,35,36].

Referring to the largest groups, the following indicator was proposed—the Firmicutes/Bacteroidetes (F/B) ratio. This ratio between these two phyla is already a documented indicator for diseases, e.g., obesity [37]. Studies described by, among others, Cattaneo et al. and Vogt et al. proved the existence of disproportions in the percentage of microflora. Samples taken from AD patients contained a reduced percentage of bacteria acting beneficially, while an increase in pathological bacteria was noticeable [38,39]. The proportion of the two phyla was also imbalanced, with a reduced level of Firmicutes, causing the dysbiosis.

It should be emphasized, however, that this does not mean directly calling individual types "good" or "bad", because the properties of the microbiome are determined by the proportions, diversity, and stability of the resulting network of dependencies [9,40].

Special attention should be paid to Firmicutes, the largest bacterial phylum in human microbiota. These bacteria are mostly Gram-positive with a thick peptidoglycan layer. Firmicutes have over 200 genera, such as Ruminococcus, Clostridium, Eubacterium, Lactobacillus, Faecalibacterium, Roseburia, and Mycoplasma [33,40]. A significant feature of these bacteria is the fermentation of indigestible carbohydrates (such as dietary fiber) and the production of short-chain fatty acids (SCFAs). SCFAs are a crucial part of the intestinal barrier, as they provide nutrients to epithelial cells, and can regulate as well as help the human immune system. In several studies from 2017 to 2021, a disproportion between microbiota producing SCFAs from healthy and AD patients was described. Research shows that AD patients experience a decrease in SCFA-producing species, such as Lachnospira, Ruminoclostridium 9, Clostridiaceae, Lachnospiraceae, and Ruminococcus. Additionally, it is worth adding that these varieties were also responsible for better results in various types of tests checking cognitive skills [38,40,41,42,43,44].

Staying on the subject of securing the digestive tract, one compound from SCFAs in particular deserves a broader discussion—butyrate. It is a desired compound by the colonic epithelial cells, for which it is a source of energy. Moreover, butyrate has anti-inflammatory properties and modulates the immune system [45], and here again, the same situation, patients with AD seem to have less numerous species such as Eubacterium rectale, Eubacterium eligens, and Eubacterium halli [42]. Referring to the flagship hotspot of AD pathogenesis, Aβ aggregation seems to be connected with butyrate production. Studies prove that in Aβ-positive patients, in comparison to Aβ-negative ones, a decrease in E. rectale species is observed [39].

Another species to be described is Lactobacillus, as it also produces valuable particles, e.g., gamma-aminobutyric acid (GABA) and acetylcholine. These structures play a significant role in neural communication, propagating specific information in the system. There are more and more studies on the use of therapy using probiotics to improve cognitive functions [46]. However, a paragraph focused directly on the applications of these studies, on the sublimation of probiotics, and the possibilities of therapy will be presented in a later part of the publication.

When it comes to Bacteroidetes, these bacteria have a Gram-negative cell wall structure, with an outer membrane built from lipopolysaccharide (LPS), which in turn is associated with the host's immune responses. These bacteria include the genera Bacteroides, Prevotella, and Xylanibacter [33,40]. Many species within this phylum stimulate intestinal function, produce valuable metabolic compounds, and support the immune system by preventing colonization of hostile pathogens [40]. Although there are also species that produce enterotoxins that disrupt the intestinal barrier. Studies also indicate a dangerous phenomenon where LPS, crossing the intestinal barrier, not only causes endotoxemia but also accumulates together with Aβ plaques in the brain [47]. If this condition is prolonged and exposure to LPS is constant, production and aggregation of Aβ progress noticeably. Moreover, there are studies suggesting a protective function of Aβ against microorganisms such as bacteria or fungi [29,48,49]. The dual phenomenon of protection and destruction of the molecule described here is not an isolated phenomenon in biological pathways; similar relationships occur in the case of other antimicrobial peptides. It is also worth noting that the studies on this phylum are not so clear. The studied relationships in the amount of this microflora in patients with AD and healthy patients are not consistent; the values are sometimes increased in some studies and sometimes decreased [38,39,41,42,44].

To summarize the above two paragraphs, a table with the most important information is presented.

The already mentioned dysbiosis contributes to the overall proinflammatory effect. The disturbance of microbiological balance affects the emerging inflammatory metabolites, leading to a series of biochemical reactions that ultimately also affect the neuroinflammation process. This comes down to the worsening of the formation and accumulation of Aβ, already described in the pathogenesis of AD.

Bacteria can be divided into those with a resultant pro- or anti-inflammatory effect. This division is determined by the relationship on the gut–brain axis, but also generally, as the metabolites produced by bacteria, which are sometimes toxins, get entangled in the biochemical pathways of the whole human body [8]. In the following part of the work, there is also a table (Table 1) summarizing several such properties of the resultant bacteria. However, it is also worth taking a closer look at them.

Studies of this type are based, for example, on measuring the number of specific bacteria in the patient's stool/ transgenic mice and the level of expression of pro-inflammatory (e.g., interleukins (IL-6, IL-18, IL-8), inflammasome complex (NLRP3), tumor necrosis factor-alpha (TNF-α)) and anti-inflammatory cytokines (e.g., IL-4, IL-10, IL-13). Such results are applied to patients with AD and healthy individuals in search of relationships [38,39,44]. And so, for example, in research conducted by Cattaneo et al., cognitively impaired patients indicate higher levels of proinflammatory cytokines (IL-6, CXCL2, NLRP3, and IL-1β). Additionally, the reduction in anti-inflammatory cytokine IL-10 was observed in this group. Their patients showed lower abundance of E. rectale and higher abundance of Escherichia/Shigella compared with both healthy controls [39].

The beneficial role of Firmicutes in the production of SCFAs has already been described earlier as an example of an anti-inflammatory effect. On the other hand, a relationship of polyunsaturated fatty acids (PUFA) and the gut microbiota with Bacteroides enrichment collected from AD patients was described. These bacteria seem to activate a pro-inflammatory PUFA cascade in the brain. The resulting PUFAs appear to stimulate microglial maturation in the brain, pathological processes associated with AD, and cognitive deficits [50].

With the growing interest in this topic, more and more studies have been conducted around this content, some based on mouse models, and some on the human microbiome. Based on the work available in the databases, a summary of several bacteria and their resulting action in the gut–brain axis is presented, with the focus on the type of inflammatory response (Table 2).

Remaining on the topic of the intestines, the bacteria that live there give their surroundings signs of inflammation, which in turn is related to macrophage dysfunction and the so-called "leaky gut" hypothesis. The resulting systemic inflammation affects the already known and well-presented above the BBB. Overall, a predominance of pro-inflammatory microbiome contributes to increased intestinal permeability, which in turn affects the strength and integrity of the BBB, contributing to the development of disorders occurring in AD [29,53].

Finally, it is also worth noting another idea describing the relationship between AD and microbiota. Some studies and theories consider the direct process of bacterial infection as a causative factor. In these connections, researchers refer to typical infectious phenomena and the possibility of spreading the pathogenic process after entering the bloodstream. Such phenomena are known, for example, in the case of viruses (particularly HSV-1, EBV, and HCMV). However, it is unclear if these events simply coexist in time without such a connection. It is difficult to prove a finding of a specific infection that could cause a disease as slowly developing as AD, with amyloid deposition that begins 15–20 years before symptoms appear [54,55].

The theory of the direct influence of microorganisms is already 30 years old. However, every few years, the potential value of this theory is regained attention again, as, among others, thanks to the discovery of antimicrobial properties of Aβ against bacteria and fungi [49]. As an example, it is worth mentioning the study conducted using the HSV-1 virus, where the Aβ oligomers seem to inhibit HSV-1 infection in vitro. This study could suggest that the encapsulation of the brain tissue with Aβ aggregates is initially a protective function before it turns into a pathogenic process through malfunctioning repair mechanisms. Although this remains speculative and requires further mechanistic validation in a clinical context [56]. These studies definitely require more recent research with clinical trials, but it is worth being aware of them. Finally, it is worth mentioning that the bacteria in the intestines are themselves a huge source of amyloids. In bacteria, e.g., E. coli, it has a protective function and helps build a biofilm that wards off the immune system's response. The structure of bacterial amyloids is indeed different from human amyloids, but there is a noticeable similarity in their tertiary structure, which triggers a similar response of the immune system to both of these formats [57,58]. Additionally, bacterial amyloids can act through molecular mimicry and affect host proteins, causing pathological cross-reactions, changing the structures of other proteins [59].

To sum this all up, with more and more information and awareness of all the connections, it seems that the right answer to the link between AD and human microbiota cannot be a single finding. It is probably the result of many individual processes, including both inflammatory processes and the direct impact of microorganisms, overlapping and interacting, some with greater force, others with less.

Studies have shown a causal relationship between the Western diet and pathological brain aging, with evidence of poorer cognitive function among older adults [78]. The Western diet, characterized by high intake of refined products (grains, sugars, saturated fats, salt, and highly processed foods) as well as extremely low fruit and vegetable consumption, resulting in very low fiber levels, has been linked to disturbances in the gut microbiome, resulting in immune dysfunction and inflammation [79,80]. For example, a Western diet rich in saturated fats (SFA) can induce the development of inflammation. SFA triggers TLR4 through CD14/MD2 activation, thereby promoting the expression of the transcription factor NF-κB, which plays a key role in the induction of pro-inflammatory mediators COX2, TNFα, IL-1β, IL-6, CXCL8, IL-12, and IFNγ. Western dietary patterns also promote hypercholesterolemia and insulin resistance, which impair blood flow in the brain and are associated with the development of both Alzheimer's disease and ADRD [81].

The Mediterranean diet (MeDi), comprising fresh vegetables, fruits, fish, legumes, cereals, and virgin olive oil, has been widely studied for its cognitive benefits. Early studies found that individuals with higher adherence to the Mediterranean diet have a lower likelihood of developing Alzheimer's disease over a follow-up period of four years, and MeDi is connected to lower mortality among AD patients [82,83]. Furthermore, a systematic review and meta-analysis by Hill et al. found that adherence to a Mediterranean-style diet was correlated with a decrease in levels of AD-related biomarkers, including β-amyloid and tau tangles [84]. Oleuropein, an antioxidant component of olive oil, has been shown to aid in the cleavage of amyloid precursor protein, a precursor of Aβ, and the aglycone compound of oleuropein has therapeutic potential in AD pathology [85].

The benefits of MeDi are a consequence of the components found in its staple products, which are summarized in Table 5.

The benefits of MeDi may be due to the components contained in its key products. Extra virgin olive oil (EVOO), particularly rich in phenolic compounds, has strong antioxidant properties [86]. EVOO metabolites are involved in microbial immunomodulation—a study by Martín-Peláez et al. showed that EVOO consumption stimulates the immune system (measured, among other things, by the level of IgA-coated bacteria in feces and CRP in plasma) [87]. EVOO consumption also enhanced the anti-inflammatory effect of the diet by reducing markers of inflammation and oxidative stress (decrease in MPO, 8-OHdG, TNF-α, and IL-6) and increasing the level of adiponectin, a protein that regulates the expression of the IL-10 gene, which supports the production of anti-inflammatory cytokines [89].

High intake of soluble fiber, abundant in MeDi products, is associated with a lower risk of AD in older adults. Fiber regulates the production of SCFAs by the gut microbiota. In a study by Cuervo-Zanatta et al., increased soluble fiber intake in APP/PS1 transgenic mice restored the balance of the gut microbiota, lowering propionate levels and increasing butyrate production, which led to reduced astrocyte activation due to increased expression of neurotrophic factors (BDNF, GDNF) and decreased expression of A1 markers (C3, Serping1). Butyrate supports the transition to the A2 phenotype—anti-inflammatory microglia [89,90].

Patients following a Mediterranean diet were found to have a greater abundance of species, particularly the Prevotella enterotype [91,92]. A study by Gutiérrez-Díaz et al. showed that MeDi increased the amount of bacteria from the Bacteroidetes, Prevotellaceae, and Prevotella groups and decreased the amount of Firmicutes and Lachnospiraceae [93].Another study showed that people following the MeDi diet had increased levels of Clostridium XIVa and Faecalibacterium prausnitzii, a known producer of anti-inflammatory butyrate: it inhibits NF-κB activation, thereby reducing the production of inflammatory cytokines, inhibits HDAC, affects the epigenetic silencing of inflammatory genes, and protects against the translocation of bacteria and toxins (e.g., LPS) into the blood, which could cause systemic inflammation [94,95,96,97].

It has been suggested that modified versions of the MeDi diet may be more effective than direct supplementation. The MIND (Mediterranean-DASH Intervention for Neurodegenerative Delay) diet, which combines elements of MeDi and DASH (Dietary Approaches to Stop Hypertension), shows even stronger links to reducing cognitive decline [98]. Based on similar components as MeDi, the MIND diet additionally emphasizes regular consumption of leafy vegetables, nuts, and berries due to their neuroprotective properties [99]. A prospective study of the MIND diet conducted by Morris et al. showed that high adherence to this diet for 4.5 years could reduce the risk of developing AD by as much as 53% [100]. However, some studies do not highlight significant benefits of this diet in preventing neurodegeneration compared to calorie restriction alone, so further research is needed [101].

One possible approach to modulating brain function and providing neuroprotection against the onset and progression of Alzheimer's disease is to directly influence the gut microbiome and its immunomodulatory activity through the use of probiotics. Probiotics, i.e., live microorganisms that have a direct beneficial effect on health, have gained interest in recent years as potential therapeutic agents in the treatment of AD, with particular emphasis on strains of the genus Lactobacillus and Bifidobacterium.

In a study by Bonfili et al., administration of SLAB51 (a probiotic formulation consisting of nine live bacterial strains: Streptococcus thermophilus, Bifidobacterium longum, B. breve, B. infantis, Lactobacillus acidophilus, L. plantarum, L. paracasei, L. delbrueckii subsp. bulgaricus, L. brevis) to mice with early-stage AD resulted in a partial improvement in the functioning of two disturbed mechanisms of neuronal proteostasis: the ubiquitin-proteasome system and autophagy, among others, through the activation of sirtuin-1 (SIRT-1)—a neuroprotective deacetylase enzyme that reduces ROS in the CNS [15].

Similar conclusions were reached by Kobayashi et al. (using the Bifidobacterium breve A1 strain in mice with AD) [102] and Medeiros et al., who supplemented transgenic 3xTg-AD mice with a cocktail containing Lactobacillus plantarum KY1032 and Lactobacillus curvatus HY7601 [103]. Probiotic intake led to a significant increase in the number of Bacteroidetes bacteria in the intestinal microflora, a decrease in astrocyte and microglia density in the entorhinal cortex and hippocampus, and an increase in the number of neurons in the hippocampus, indicating a neuroprotective effect [103].

Probiotics may also contribute to slowing the progression of cognitive impairment and Alzheimer's disease pathology by strengthening the integrity of the gut and BBB while alleviating inflammation in the gastrointestinal tract, systemic circulation, and central nervous system. A decrease in plasma inflammatory markers such as IL-6 and TNF-α has been observed in individuals treated with a probiotic cocktail, while mediators such as TNF-α are responsible for suppressing the expression of tight junction proteins in the brain and intestine, hence greater integrity of, among others, the BBB [104].

Prebiotics, a group of beneficial nutrients broken down by the gut microbiota, have been the subject of growing interest in recent years as potential factors in preventing Alzheimer's disease (AD) and delaying its progression [105]. The most commonly used prebiotics include fructooligosaccharides (FOS), galactooligosaccharides (GOS), and trans-galactooligosaccharides (TOS), and their fermentation by gut bacteria leads to the formation of SCFAs, such as lactic, butyric, and propionic acids [106].

FOS are a frequent subject of research due to their promising therapeutic potential in AD via the gut–brain axis. In a study by Sun et al., six weeks of oral administration of FOS to APP/PSEN1dE9 transgenic mice improved cognitive function and reduced AD-related pathological changes by increasing the expression of synapsin I and postsynaptic density protein 95 (PSD-95). In addition, FOS treatment increased GLP-1 levels and decreased the expression of its receptor GLP-1R, thereby modulating the gut microbiota-GLP-1/GLP-1R signaling pathway [107]. In another study, FOS not only improved oxidative stress and inflammation markers but also regulated neurotransmitter synthesis and secretion. Histological analysis also showed reduced neuronal apoptosis and decreased levels of key AD markers such as Tau and Aβ1-42 [108].

GOS, oligosaccharides, also exhibit neuroprotective effects. In a study by de Paiva et al., administration of FOS and GOS to mice on a high-fat diet led to a reduction in neuroinflammation and neuronal damage (as demonstrated by lower levels of TNF-α, COX-2, caspase-3, Iba-1 cells, and GFAP). The treatment improved synaptic plasticity, restored learning and memory abilities, and supported the insulin signaling pathway through activation of the IRS/PI3K/AKT pathway. In addition, it reduced amyloid β formation and Tau phosphorylation, helped restore the balance of the gut microbiota—particularly by increasing the abundance of Bacteroidetes bacteria—and reduced intestinal inflammation and permeability [109].

Xylooligosaccharides (XOS) are indigestible oligosaccharides with high prebiotic potential [110]. They strongly stimulate the growth and diversity of Bifidobactera in the gut [111]. XOS supplementation also selectively promotes the expansion of strains Bifidobacterium adolescentis, known for its involvement in modulating neuroinflammatory pathways [112].

Increasing the diversity of the intestinal microflora through XOS supplementation reduced intestinal inflammation by lowering the levels of pro-inflammatory cytokines IL-1β and IL-6 and immunoregulatory IL-10. Additionally, it increased the expression of tight junction proteins in the intestine, contributing to the restoration of both the epithelial barrier and the blood–brain barrier [113].

Probiotics and prebiotics appear to be promising therapeutic agents for the treatment of AD; however, the currently available scientific evidence comes mainly from studies in animal models. Therefore, further clinical studies involving humans are necessary to confirm their efficacy and practical applicability.