A Comprehensive Review of the Role of the Microbiota–Gut–Brain Axis via Neuroinflammation: Advances and Therapeutic Implications for Ischemic Stroke

The concept of the MGBA was first introduced by Sudo et al. in 2004 [], a communication network encompassing the gut, nervous system (central, autonomic, and enteric), HPA (hypothalamic–pituitary–adrenal) axis, the neuroendocrine system, the immune system, gut microbiota, intestinal mucosal barrier, and the BBB []. This concept underlies the dynamic bidirectional communications that exist between the CNS and the GIT, functioning through three main pathways: immune, neuronal (including the sympathetic, parasympathetic, and enteric nervous systems), and endocrine (involving the HPA axis) []. These pathways facilitate dynamic interactions among gut microbiota, gut metabolism, and the CNS, ultimately influencing brain function (). A crucial aspect of this axis is the bidirectional communication system that links the gut microbiota and the brain. The CNS transmits signals to the gut, regulating functions such as peristalsis, mucus secretion, and the mucosal immune system, influencing the diversity of gut bacteria through the ANS (autonomic nervous system). On the other hand, gut bacteria communicate with the CNS and immune system by releasing neurotransmitters, metabolites, and immune mediators, which can either ameliorate or exacerbate brain-related conditions [,]. ^{10 11 12} Figure 1 ^{13 14}

There are unique microbiomes in almost every ecological niche of our body, with the major sites of colonization being the skin, respiratory tract, genitourinary tract, eyes, and GIT []. Significant advancements in sequencing technologies and the development of microbiome bioinformatics tools have made it increasingly affordable to analyze microbiota composition. To date, 2172 species have been identified across the human bodies, including 12 different phyla, with 386 species of anaerobic bacteria thriving in the mucosal regions, such as the GIT and oral cavity []. While the oral and pulmonary microbiota are critical, the majority of our microbial residents reside in the gut, which houses a diverse array of microorganisms, including yeasts, archaea, helminths, viruses, protozoa, and bacteria—the latter has been the most extensively studied []. ^{15 16 15}

In humans, these microorganisms serve diverse functions, such as synthesizing vitamins B and K, producing derivatives such as SCFAs (short-chain fatty acids), bile acids, TMAO (trimethylamine N-oxide), LPS (lipopolysaccharides), and phenylacetylglutamine. Moreover, they are instrumental in metabolizing key compounds like amino acids (e.g., glycine, glutamic acid, Gamma-Aminobutyric Acid (GABA), aspartic acid), peptides (e.g., vasopressin, somatostatin, neurotensin), biogenic amines (e.g., norepinephrine, serotonin, dopamine), and ACh (Acetylcholine) []. They also ferment undigested carbohydrates and defend against pathogens. Ideally, the bacterial community residing in the gut should have evolved to function in a symbiotic manner with the host, facilitating digestion and fostering an appropriate immune response. ¹⁶

However, dysbiosis of the gut can lead to excessive bacterial growth in the intestinal region, resulting in an increase in the permeability barrier and the induction of systemic inflammation. This inflammatory cascade overactivates the immune system, setting the stage for the development of various diseases []. A notable clinical example of this is hepatic encephalopathy, where targeting the microbiota with antibiotics can help reduce ammonia production in the gut, a key contributor to this disease []. By altering the gut microbiome, antibiotics may improve cognitive function and overall outcomes in patients suffering from this condition, and this also synergizes with the concept of the microbiome-gut–brain axis. ^{17 18}

So far, the MGBA has been implicated in a wide range of diseases, including neurodegenerative conditions such as Alzheimer’s disease, Parkinson’s disease, and ALS (amyotrophic lateral sclerosis). It also plays a role in ischemic stroke, depression, autism spectrum disorder, dementia, mild cognitive impairment, multiple sclerosis, inflammatory bowel disease, drug-resistant epilepsy, and insomnia. These diverse conditions underscore the importance of understanding the MGBA as a potential therapeutic target for a variety of neurological and systemic disorders []. ¹⁹

When discussing immune cells within the CNS, microglia inevitably warrant particular emphasis. Microglia—the resident immune cells of the CNS—undergo dynamic functional and phenotypic transformations in response to brain injury [27]. These changes are particularly well-characterized in the Middle Cerebral Artery Occlusion (MCAO) model, which serves as the gold standard experimental paradigm for ischemic stroke research in rodents [28]. This model recapitulates the key pathophysiological features of human stroke through surgical occlusion of the middle cerebral artery (MCA), with both transient (tMCAO) and permanent (pMCAO) variants allowing for an investigation of different ischemic durations. Under homeostatic conditions, microglia perform a variety of essential physiological functions, including regulating neurogenesis and angiogenesis, maintaining BBB integrity, synaptic pruning and remodeling, facilitating synaptic transmission, supporting myelin health, and engaging in the phagocytosis and clearance of apoptotic neurons and debris [29]. Notably, a recent single-cell RNA sequencing study revealed two distinct microglial subclusters that exhibit differential activation patterns under MCAO-induced ischemic conditions [30]. These two subclusters located in different regions of the ischemic brain: ICAM (ischemic core-associated microglia) in the infarcted area and IPAM (ischemic penumbra-associated microglia) in the surrounding penumbra [30]. These subclusters exhibit distinct functional and metabolic profiles, as demonstrated by GSVA (gene set variation analysis) (Figure 2). It was found that ICAM primarily relies on glycolysis, suggesting that they shift towards anaerobic metabolic pathways in response to ischemic injury. IPAM depend on the TCA (tricarboxylic acid) cycle and oxidative phosphorylation, indicative of a more oxidative metabolic profile. This metabolic divergence is linked to their distinct roles in stroke pathology. ICAM are associated with excessive pro-inflammatory responses and enhanced chemotaxis, suggesting they may contribute to tissue damage and exacerbate the acute inflammatory response during ischemic stroke. Their hyperactivation could accelerate neuronal injury and worsen stroke outcomes.

In comparison, IPAM display enriched metabolic pathways related to amino acids, lipids, and carbohydrates. Since lipid metabolites are essential components of myelin, IPAM may compensate for the loss of oligodendrocyte function by supplying lipid components. The role of lipid metabolites in neuroinflammation has been well-documented, and the anti-inflammatory properties of sphingolipid signaling are of increasing interest in neurodegenerative and neuroinflammatory research. The balance between these microglial subtypes and their metabolic pathways may therefore be a critical factor in determining stroke outcomes, influencing both the progression of neuroinflammation and the potential for recovery.

Microglial cells exhibit distinct functional states at different stages following ischemic stroke, positioning them as key players with dual roles in neuro-injury and neuroprotection (Figure 3). Early neuroinflammation, driven by microglial activation, serves as an adaptive microglial mechanism, providing neuroprotective effects by facilitating tissue repair, clearing cellular debris, and eliminating pathogens.

Following ischemic stroke, DAMPs are released, activating the PRRs on microglia. Additionally, accumulated ATP (adenosine triphosphate) and UDP (uridine triphosphate) in the ischemic tissue can activate P2 purinergic receptors on microglia. These events, coupled with the loss of immunosuppressants and chemokines from the neuronal surface, amplify microglial activation, leading to a vicious cycle of overactivation. This hyperactivity of microglia contributes to increased neuronal death and intensifies neuroinflammation, worsening the overall brain injury. In the late stage of AIS, activated microglia adopt a neuroprotective role, helping to mitigate excessive inflammation and restore homeostasis within the brain. Also, as the primary phagocytic cells in brain tissue, microglia can quickly clear cellular debris, reduce DAMPs, and protect the brain from secondary inflammatory response damage, promoting the recovery of nerve function and the re-establishment of the neural network.

Upon the onset of an ischemic stroke, quiescent microglia are rapidly activated and move towards the M1 subtype, initiating an inflammatory cascade characterized by the secretion of various cytokines and chemokines. This M1-driven inflammatory response disrupts the BBB and contributes to neuronal death. Conversely, microglia can also shift to the M2 subtype, which exhibits neuroprotective properties and secretes anti-inflammatory factors to facilitate tissue regeneration and repair [31]. The M1 subtype has been implicated in the generation of Th1 cells, with M1-polarized macrophages mediating the differentiation of helper T cells into Th1 and Th17 subsets. Given its pro-inflammatory nature, the M1 subtype can replace the M2 subtype, thereby perpetuating the inflammatory cycle.

The expression of major histocompatibility MHC-II (complex class II), along with CD16/32, CD40, and CD86 on the surface of M1 microglia, further stimulates the production of pro-inflammatory cytokines, such as TNF-α, IL-1β, IL-6, IL-12, and IL-23. Furthermore, microglia gradually lose their homeostatic signatures and become activated with age or in pathological conditions, adopting disease-associated phenotypes that release pro-inflammatory cytokines and chemokines [32]. Several genetically distinct microglial subtypes have been identified, including homeostatic microglia and “disease-associated microglia” (DAM) or the “microglial neurodegenerative phenotype”. Therefore, targeting the M1/M2 phenotypic switch of microglia or the DAM and the microglial neurodegenerative phenotype in the post-stroke period holds great promise as a therapeutic strategy to modulate neuroinflammation and promote recovery [33].

Given this, numerous clinical and laboratory studies have focused on exploring targets for intervention in microglial differentiation. Emerging evidence indicates that different dietary fibers can significantly modify the gut microbiome’s composition and function in obese mice on a high-fat diet [34,35]. This intervention is crucial for improving microglial cell maturation, which is essential for optimal brain function and health [36]. In addition to regulating microglial homeostasis, metabolites from gut microbes play a crucial role in triggering microglial cell death. Elevated concentrations of metabolites produced by gut microbes can cross the BBB, triggering programmed cell death in microglial cells. This process, which becomes more pronounced with advancing age, may contribute to the neuroinflammatory milieu observed in neurodegenerative diseases.

Among the gut-derived metabolites, SCFAs warrant particular attention due to their indispensable role in regulating microglial differentiation. SCFAs are important metabolic by-products of fiber digestion by gut bacteria, including acetate, propionate, and butyrate, which influence microglial behavior and function, ensuring proper development and activation in response to various signals from the body. The ability of the gut microbiome to influence microglial activation highlights the importance of the gut–brain axis in maintaining neural health. In certain pathological conditions, impaired microglial phagocytosis can result in the accumulation of toxic compounds, and excessive microglial activity may lead to neuronal loss and neurodegeneration through the phagocytosis of neurons [37].

In MCAO mouse models, supplementation with SCFAs significantly improved the recovery of motor function in the affected limb [38]. This improvement is believed to be mediated by the modulation of microglial activity. Following ischemic stroke, the intestinal flora activates microglia by producing endogenous AHR (aryl hydrocarbon receptor) ligands and SCFAs, which exert neuroprotective effects. The gut flora metabolizes tryptophan into AHR ligand-forming metabolites, which modulate microglial activation in neuroinflammatory conditions through their action on the microglial AHR [39,40]. This signaling pathway plays a pivotal role in the neuroprotective effects observed in the context of stroke and offers a potential therapeutic avenue for mitigating neuroinflammation and promoting recovery after ischemic events.

While gut-derived metabolites (e.g., SCFAs) have been the focus, emerging evidence underscores the indispensable roles of ApoE (Apolipoprotein E) and T cells in modulating microglial differentiation. Recent findings have clarified the complex interactions among microglia, ApoE, and T cells in neurological diseases [41]. ApoE, a lipid and cholesterol transporter, plays multiple roles in the CNS, including the regulation of microglial and astrocytic function, the maintenance of cerebrovascular and BBB integrity, myelin dynamics, and the modulation of neuronal network activity. Furthermore, T-cell infiltration into the CNS is critical for the activation of microglia and astrocytes in animal models of ALS [42]. In these models, treatment with natalizumab, which reduces immune cell recruitment, resulted in a significant reduction in pathological changes within the CNS, including decreased motor neuron degeneration, delayed onset of paralysis, and increased survival. These findings highlight the potential therapeutic benefit of targeting the signaling pathways between microglia and T cells to modulate neuroinflammation and improve outcomes in neurodegenerative diseases.

The GIT is a key immune organ in the body, containing approximately 70% of immune cells [43], including a diverse array of immune cell types, such as B cells, Treg (regulatory T cells), macrophages, dendritic cells, γδT cells, αβT cells, and others. The intestinal mucosal immune system can be divided into three parts involved in defense: the columnar epithelial cell layer, the lamina propria, and the gut-associated lymphoid tissue. The intestinal mucosal immune defense comprises two distinct categories of sites: the immune effector sites and the immune inductive sites [43]. These sites collectively regulate a vast array of gut bacteria, maintaining a balance between tolerance of commensal microorganisms and effective immune regulation [44].

Inflammation and immunity play a crucial role in the pathophysiology of ischemic stroke, influencing all stages of the disease, from the development of risk factors to neurotoxicity and tissue remodeling during recovery [45,46]. Experimental research using rodent models has underscored the critical role of the immune system in stroke, particularly the brain-infiltrating lymphocytes originating from the intestinal immune compartment. These lymphocytes facilitate communication along the gut–brain axis, a key process in the neuroinflammatory response following ischemic injury [47,48,49,50].

The neuroinflammatory response in the brain is triggered and sustained by various components of the ischemic injury process, including necrotic cells, debris, and ROS (reactive oxygen species) [51]. This response involves the activation of resident inflammatory cells, the release of inflammatory mediators, and the migration of leukocytes across the BBB [52]. This, in turn, activates T and B lymphocytes, initiating an adaptive immune response [53]. In vivo cell-tracking studies, such as those conducted by V. Singh, have demonstrated the migration of intestinal lymphocytes to the ischemic brain, highlighting the integral role of the gut in modulating the immune response in stroke [54].

The neuroinflammatory cascade following ischemic injury is initiated by necrotic and apoptotic cells releasing DAMPs (e.g., HMGB1, ATP), which activate microglia through TLR4 (Toll-like receptor 4)/NLRP3 pathways [46]. This process is exacerbated by ROS: while the ischemic core shows sustained, high-level ROS (O₂⁻, H₂O₂) due to mitochondrial collapse (Complex I/III inhibition and mPTP opening), the penumbra exhibits moderate, partially reversible ROS increases, mediated by the Nrf2/ARE antioxidant response [45,55]. In reaction to ischemic injury, necrotic cells, debris, and ROS participate in a second phase of immune activation—neuroinflammation, during which the neutrophils, microglia or macrophages, mast cells, lymphocytes, and NK T-cells are mobilized to the site of injury to initiate the neuroimmune response [56]. The activation of PRRs on these immune cells triggers the production of pro-inflammatory cytokines, such as IL-1β and TNF-α, which create an inflammatory environment, characterized by an increase in IL-17, perforin, granzyme and reactive oxidants, contributing to the worsening of tissue damage [57].

The necrotic tissues and brain cells release CNS-specific cryptic antigens, which can be recognized by APCs (antigen-presenting cells) in the brain. These APCs then migrate through the meningeal lymphatics into the peripheral circulation, eventually reaching the lymphoid organs, where they promote the differentiation of T and B cells. These differentiated self-reactive lymphocytes return to the brain tissue, potentially contributing to post-stroke sequelae, including chronic neuroinflammation and further neuronal injury [58].

T cell subsets play pivotal roles in this process, including both helper T cells (Th) and Treg. Both CD4+ and CD8+ T cell counts significantly increase after stroke, correlating with poorer functional outcomes. CD4+ T cells play a crucial role in the adaptive immune response within the intestine and can differentiate into various subtypes, including Th1, Th2, Th17 cells, and Tregs. Pro-inflammatory subsets, specifically Th1 and Th17 cells, can promote neuroinflammation [53,59]. Th1 cells exacerbate neuroinflammation and the activation of microglia by secreting cytokines such as IL-2, IL-12, and IFN-γ (Zhang et al., 2024 [59]). Th17 cells, on the other hand, can activate matrix metalloproteinases and contribute to the destruction of the BBB structure by producing cytokines such as IL-17A, IL-17F, and IL-22 [2,53,60,61].

Notably, the cytokine profiles also differ regionally (Table 2): the core is dominated by pro-inflammatory TNF-α and IL-1β from dying neurons and activated microglia, whereas the penumbra shows mixed signals, including anti-inflammatory IL-10 and TGF-β from infiltrating Tregs. Mitochondrial dysfunction plays a central role, with macrophages’ depolarization impairing ATP synthesis (leading to excitotoxicity) and released mtDNA fueling cGAS-STING-mediated neuroinflammation [62].

The IEB (intestinal endothelial barrier) is essential for maintaining intestinal homeostasis, serving as a physical barrier and a coordinator of immune responses. It comprises epithelial cells, goblet cells, Paneth cells, and enterochromaffin cells. Communication between the intestinal microbiome and peripheral nerve cells within the gut–brain axis occurs through three interrelated pathways. Endothelial cells in the gut respond to signaling molecules released by gut bacteria by initiating the release of neuropeptides. The receptors on EECs (enteric epithelial cells) are specifically located on vagal sensory neurons that project into the gut, influencing various physiological functions associated with digestion and metabolic regulation [152,153,154,155].

Moreover, evidence suggests that an individual’s age can affect the integrity of the IEB following ischemic stroke. With aging, the gut barrier weakens, allowing pro-inflammatory substances from gut microbes and harmful bacteria to enter the bloodstream, leading to systemic inflammation and neuroinflammation [156]. In vivo studies in animal models have shown that stroke significantly disrupts intestinal homeostasis, with older mice exhibiting more pronounced effects compared to younger ones, suggesting that age may be a critical factor in the extent of post-stroke gut barrier disruption [157].

The current evidence suggests that stroke can lead to a disruption in the integrity of the IEB, resulting in the deterioration of the intestinal villous epithelium, increased permeability, and compromised tight junctions, which facilitates the leakage of harmful substances into the bloodstream, leading to a reduction in mucus production, further weakening the intestinal barrier and increasing susceptibility to enterogenic sepsis [158]. The majority of γδT cells in the human body are located on the surface of the intestinal epithelium, where they play a role in the innate immune response of the intestine. Following ischemic stroke, γδT cells migrate from the gut through the peripheral circulation to the brain membrane and secrete IL-17 into the damaged brain tissue, inducing the production of increased chemokines in the brain parenchyma. This eventually leads to a massive influx of neutrophils and exacerbates ischemic neuroinflammation [159].

Many gut microbes and their byproducts can influence the outcome of a stroke by modulating the intestinal mucosal barrier. A high fiber intake has been shown to promote the growth of fiber metabolizers and SCFA producers, which can enhance mucus secretion and maintain a healthy mucus layer [160,161]. On the other hand, low-fiber diets in mice led to an increase in the population of mucus-degrading bacteria, with a thinner mucus layer and an elevated risk of infection [162]. Other metabolites involved in maintaining intestinal barrier integrity include indole and its derivatives. Compounds such as indole-3-ethanol, indole-3-pyruvate, and indole-3-aldehyde reinforce the apical junctional complex, thereby enhancing the resilience of the intestinal barrier [163]. Furthermore, these indoles can inhibit microglial and NLRP3 inflammasome activation by engaging the microglial AHR, mitigating post-stroke neuroinflammation [164].

The BBB is a sophisticated physiological and biochemical barrier that maintains brain homeostasis by regulating the exchange of substances between the brain and its external environment. It is responsible for removing harmful metabolic byproducts and internal endotoxins from the brain [165]. The integrity of the BBB is maintained by tightly joined, non-porous endothelial cells connected by tight and adherent junctions. These endothelial cells share a basement membrane with pericytes and astrocytes, along with neurons and microglia, forming neurovascular units that are crucial for maintaining BBB integrity. Inflammatory molecules such as IL-6 and IL-8 are believed to disrupt neurovascular function and may contribute to the increased vascular permeability observed in the BBB during inflammation [166,167]. These disruptions facilitate the infiltration of peripheral immune cells and inflammatory mediators into the CNS, exacerbating neuroinflammation and contributing to the pathogenesis of neurological disorders.

Numerous studies suggest that metabolites produced by the gut microbiota significantly influence the regulation of BBB integrity [168,169,170,171,172]. SCFAs, among the most-studied microbial metabolites, have a notable impact on BBB integrity. Research using in vitro BBB models has revealed a protective effect of SCFAs on maintaining the barrier.

The role of TMAO in BBB function is more variable. Some studies link TMAO to neuroinflammation due to its ability to activate microglia and astrocytes. TMAO has also been associated with negative impacts on neuronal health, including neuronal aging, oxidative stress, and alterations in synaptic plasticity [89,173,174,175,176,177,178,179]. These findings suggest that elevated TMAO levels may exacerbate the inflammatory processes underlying neurodegenerative diseases. However, it has also been found that the chronic administration of low doses of TMAO can protect against LPS-induced BBB damage and memory deficits in C57Bl/6J mice [168]. This raises the possibility that TMAO’s effects on the BBB may depend on dose and the specific context of its administration.

Inflammation is known to have an adverse impact on BBB function [167]. Pro-inflammatory substances have the potential to compromise the integrity of the BBB, which could allow substances from the bloodstream to enter the brain. This breach in the BBB could establish a connection between the CNS and the peripheral immune system, potentially leading to neuroinflammatory conditions.

Diet, as a modifiable factor, exerts an influence on the gut microbiota across both short- and long-term periods. Several studies proved that vegetable consumption may be associated with a reduced risk of stroke [181,182,183]. A large European cohort study revealed that a higher intake of fruits, vegetables, fiber, and dairy products was linked to a lower risk of ischemic stroke, while a higher consumption of red meat was associated with an increased risk. Additionally, higher egg intake was found to correlate with an elevated risk of hemorrhagic stroke [181]. Similarly, a Taiwanese case–control study found that adhering to a vegetarian diet in Taiwan was associated with a lower risk of both ischemic and hemorrhagic stroke [182].

In 2023, a case–control study among the Lebanese population investigated the impact of the DASH (Dietary Approaches to Stop Hypertension) diet on ischemic stroke and found that the diet was protective against such strokes and associated with less disability [184]. The DASH diet emphasizes the consumption of vegetables, fruits, whole grains, low-fat dairy, lean meats, and nuts, along with a significant reduction in sodium intake [185]. In contrast, a recent study indicated that a plant-based diet might actually increase the risk of stroke compared to red meat consumption, suggesting that an exclusively plant-based diet may not always confer the expected health benefits [186].

The ketogenic diet, which is very high in fat and virtually devoid of carbohydrates, has yielded conflicting results regarding its impact on gut microbiota. While some research indicates that such diets may disrupt the balance of the gut bacteria and lead to inflammation [187,188], others have highlighted the potential benefits of ketogenic diets in the context of stroke-related diseases, [189,190] as diet-induced ketosis may have a positive relation with the development of an anti-inflammatory microglial phenotype [189].

The relationship between coffee consumption and stroke risk remains contentious. A study [191] involving 13,462 stroke cases and 13,488 controls from the INTERSTROKE database found that high coffee intake was associated with an increased stroke risk, whereas tea consumption was linked to a reduced risk. In contrast, a cohort study using UK Biobank data [192] suggested that drinking coffee and tea, either separately or together, was associated with a lower risk of stroke and dementia. Another interesting study found that carbonated beverages were linked to higher odds of ischemic stroke and intracerebral hemorrhage, while high water intake correlated with lower ischemic stroke odds, with notable regional variations [193]. To date, only a fraction of research has explored the interplay between diet, stroke, and gut microbiota. A comprehensive investigation into dietary impacts on gut microbiota could offer novel insights in the prediction of stroke outcomes (Table 3).

In summary, dietary interventions targeting post-stroke neuroinflammation and the subsequent modulation of neurological outcomes through the MGBA still face several practical limitations. For instance, the precise dose–response relationship between dietary fiber intake and stroke incidence or severity remains unclear. Additionally, significant inter-individual variability—such as differences in baseline gut microbiota composition, prior antibiotic use, and long-term dietary patterns before stroke onset—may introduce bias into the observed outcomes.

Antibiotic treatment can significantly alter the composition and function of the intestinal microbiota, potentially disrupting the microbiome’s homeostasis and influencing stroke outcomes [194]. The gut microbiota plays a crucial role in maintaining immune regulation, metabolic balance, and neurological function. When antibiotics disturb this delicate ecosystem, dysbiosis can occur, leading to a reduction in beneficial bacteria (e.g., Lactobacillus and Bifidobacterium) and an overgrowth of pathogenic or opportunistic microbes [195]. This imbalance may compromise gut barrier integrity, increase systemic inflammation, and modulate immune responses, all of which can impact stroke pathogenesis and recovery [196]. Interestingly, while antibiotic-induced microbiota disruption is generally detrimental, some studies suggest that the targeted modulation of gut bacteria may have therapeutic potential in stroke. Utilizing antibiotics to target gut microbiota can lead to the production of inflammatory factors such as IL-1, IL-6, and iNOS, which may reduce the formation of infarcted areas. As mentioned in Section 3.2 [48], Benakis C. et al. conducted animal experiments that demonstrated that gut microbial changes due to antibiotic treatment following ischemic stroke can result in modifications to dendritic cells. These alterations disrupt intestinal immune homeostasis, leading to an increase in regulatory T cells and a decrease in IL-17-producing T cells, mitigating ischemic brain damage in MCAO mice. Importantly, whether neuroprotection is mediated by gut microbiota is dependent on the balance between IL-10 and IL-17, suggesting an intricate connection between gut microbiota and the immune system. However, the use of broad-spectrum antibiotics, which reduce gut microbial populations, does not affect brain damage within the first day post-stroke, but may suppress systemic immunity, leading to increased mortality between days 5 and 7 following stroke [197]. These findings highlight that the role of antibiotics in preventing and treating stroke remains a topic of debate.

Probiotics are defined as “live microorganisms that, when administered in sufficient quantities, confer health benefits on the host.” While prebiotics are “substrates selectively utilized by host microorganisms to promote health”. From a microbiological perspective, prebiotics are non-digestible dietary compounds that selectively stimulate the growth and/or activity of beneficial gut microbiota, thereby conferring indirect health benefits. In contrast, probiotics are live microorganisms which, when administered in adequate amounts, directly confer a health benefit to the host by modulating gut microbial composition and function. Both probiotics and prebiotics are effective in restoring beneficial gut microbiota and metabolic functions, enhancing the production of SCFAs and thus improving the integrity of biological barriers and reducing systemic LPS levels. As a result, systemic inflammation and glial activation decrease, which mitigates neurodegenerative disease pathology.

Prebiotics, which are non-digestible compounds found in high-fiber foods like garlic, onions, bananas, asparagus, oats, and chicory root, exert beneficial effects on the host by influencing the composition and activity of the intestinal microbiota through microbial metabolism [198]. Commonly studied prebiotics include inulin, fructooligosaccharides (FOS), and galactooligosaccharides (GOS), which promote the growth of Lactobacillus and Bifidobacterium species [199,200,201,202,203].

The administration of probiotics, which are common in fermented foods like yogurt, kefir, sauerkraut, kimchi, miso, and supplements, has been shown to restore intestinal microbial balance, reduce the production of TMAO, increase SCFA production, alleviate damage to tight-junction proteins, and decrease both adaptive and innate immune activation, thus minimizing post-stroke inflammatory damage [204,205,206]. A higher risk of stroke has been associated with an imbalance in the gut microbiota, characterized by an increase in opportunistic pathogens and lactate-producing bacteria and a decrease in butyrate-producing bacteria [207]. A combination of early enteral nutrition and probiotics can effectively improve the nutritional status of stroke patients, regulate intestinal flora and mucosal barrier function, and enhance immune function, helping to reduce the incidence of infectious complications and gastrointestinal motility disorders. This therapeutic approach also contributes to faster recovery and better outcomes for stroke patients [208,209]. Additionally, probiotic treatment can protect gut barrier integrity and increase the expression of brain GLP-1 receptors and the secretion of gut GLP-1, a hormone that regulates 5-HT levels and appetite [15].

Despite their promising benefits, the use of probiotics to prevent and treat strokes caused by gut bacteria remains a conflict. Some probiotic strains may exacerbate inflammation by promoting M1 macrophage polarization [210,211]. A randomized controlled trial involving elderly care-home residents found that probiotics had no significant effect on plasma immune mediator levels, neutrophil and monocyte phagocytosis, or blood culture responses to immune stimulation [212]. This has raised concerns regarding the varying effectiveness of probiotics across different populations and conditions. Challenges in probiotics use include their susceptibility to a low gastric pH and digestive enzymes, which can lead to inactivation and reduced bioactivity. Moreover, biases can be introduced throughout the sample processing stages, including collection protocols, preservative selection, storage temperature, DNA extraction, library preparation, sequencing, and bioinformatics analysis [213] (Table 4).

FMT has emerged as an innovative therapeutic approach, receiving attention due to its potential to modulate the gut microbiota and treat various conditions. FMT involves the transfer of stool from a healthy donor to a recipient, aiming to restore the recipient’s gut microbiota balance. This method is being explored as a treatment for neurological disorders, including stroke. To date, much of the research on FMT for stroke has been conducted in animal models. Studies have shown that FMT can exert neuroprotective effects by reducing pro-inflammatory bacteria and gut microbiota metabolites, as well as diminishing inflammatory responses and oxidative stress in the brain [214]. Research by Benakis et al. indicates that recolonizing mice with a dysbiotic microbiome can induce pro-inflammatory T-cell polarization in both the intestinal immune compartment and the ischemic brain. Additionally, therapeutic FMT has been shown to normalize brain lesion-induced dysbiosis and improve stroke outcomes in animal models [48]. Furthermore, the therapeutic transplantation of fecal microbiota has been demonstrated to normalize brain lesion-induced dysbiosis and improve stroke outcome [54]. Based on these findings, it is hypothesized that gut ecological dysregulation can significantly impact neuroinflammation, metabolism, and immune homeostasis following brain injury, suggesting that FMT could be a promising strategy for mitigating these effects in stroke patients. However, further research is needed to translate these preclinical findings into clinical practice and to address potential safety and efficacy concerns.

The gut microbiota exerts profound effects on human physiology by modulating host immunity, metabolic processes, and neurological functions through specialized bacterial communities. Of particular significance are the Lactobacillus species, Gram-positive, facultative anaerobic bacteria that dominate mucosal surfaces and the intestinal tract, where probiotic strains including L. acidophilus and L. rhamnosus strengthen epithelial barrier function, regulate immunological activity, and competitively exclude pathogenic microorganisms [215]. Equally noteworthy is Akkermansia muciniphila, a mucolytic Gram-negative anaerobe that maintains intestinal homeostasis through its unique capacity to utilize mucin as its primary energy source. Its colonization has been inversely correlated with systemic inflammation and metabolic syndrome, while it is positively associated with enhanced insulin signaling [216,217]. In contrast, Escherichia coli exemplifies the dichotomous nature of gut symbionts: commensal variants participate in essential physiological processes including vitamin K biosynthesis and niche occupation, whereas enteropathogenic serotypes such as O157:H7 possess virulence factors that can precipitate life-threatening gastroenteritis, illustrating the delicate balance between microbial mutualism and pathogenicity in the gastrointestinal ecosystem [218,219]. Laboratory studies have revealed that patients with ischemic stroke often exhibit a higher prevalence of opportunistic pathogens such as Bifidobacterium, Oscillibacter, and Enterobacter, and lower levels of beneficial genera like Faecalibacterium and Bacteroides in their feces or intestinal contents compared to healthy controls [220,221,222,223]. In a case–control study conducted by Na Li, 30 cerebral ischemic stroke (CI) patients and 30 healthy controls were enrolled to compare their fecal gut microbiota profiles using Illumina sequencing of the 16S rRNA gene. The result found that CI patients had significant dysbiosis, with an enrichment of short-chain fatty acid-producing bacteria such as Odoribacter and Akkermansia [224]. However, the reliability of these findings is limited due to the small sample size and the lack of consideration of other factors that could influence gut microbiota, such as dietary habits and medication use. These results collectively suggest that the composition of the gut microbiota may contribute to an individual’s risk of stroke.

The long-term outcomes of FMT as a therapeutic intervention are not yet fully understood, and some conflicting results have been reported. For instance, Vendrik et al. observed an increase in mortality following FMT in MCAO mice models [225], questioning the safety of this therapy. Regarding the application of FMT in stroke, only a limited number of human studies have been completed or are ongoing. To clarify the role of FMT in the context of stroke, substantial, double-blind, randomized controlled trials will be necessary.