CI is closely related to changes in the composition of the intestinal microbiota ( Table 1 ). Analysis of the proportion of various components of the intestinal microbiota in patients is helpful in identifying possible symptoms of CI as early as possible. Yamashiro et al. analyzed the fecal gut microbiota composition of CI patients and control subjects. The results showed that CI was closely associated with an increase in Atopobium cluster and Lactobacillus ruminis, and a decrease in the Lactobacillus sakei subgroup (Yamashiro et al., 2017). A prospective observational study of short-chain fatty acid (SCFA) profiles in patients with ACI in China showed that SCFAs-producing bacteria (Roseburia, Bacteroides, Lachnospiraceae, Faecalibacterium, Blautia, and Anaerostipes) were less abundant in ACI patients while Lactobacillaceae, Akkermansia, Enterobacteriaceae, and Porphyromonadaceae were overgrown, which reflected the dysregulation of the intestinal microbiota in ACI patients (Tan et al., 2021). However, another study showed the opposite. The analysis of the gut microbiota of ACI patients and healthy controls by Li et al. showed an increase in SCFAs producing bacteria in patients, including Odoribacter, Akkermansia, Ruminococcaceae_UCG_005, and Victivallis (Li et al., 2019).

Atherosclerosis is a possible cause of CI. The gut microbiota analysis of patients with atherosclerotic plaque who had undergone carotid endovascular resection for mild CI, transient ischemic attack, or transient amaurosis leaurosis showed that Collinsella enriched in symptomatic atherosclerotic plaque, while Eubacterium and Roseburia decreased. Patients’ metagenome is rich in genes associated with peptidoglycan biosynthesis, suggesting that intestinal metagenomic increased peptidoglycan production may promote symptomatic atherosclerosis by primes the innate immune system and enhances neutrophilic function (Karlsson et al., 2012). Although studies have demonstrated a relationship between clot histology and CI, few cases involving Candida have been reported. Clot analysis of a patient with ACI showed a notable presence of Candida albicans (Walker et al., 2019).

The inflammatory response to sterile tissue injury is a key pathophysiology of organ-specific injury, including CI (Chamorro et al., 2012). Previous studies have shown that pro-inflammatory Th1, Th17, and γδ T cells are associated with increased inflammatory damage and poor prognosis, while Treg cells suppress neuroinflammatory responses to brain injury (Gelderblom et al., 2012; Liesz et al., 2013). The evidence suggested that the gut microbiota is a key regulator of T cell homeostasis and is closely related to the maturation of the immune system and the coexistence of the maintenance host and microbes. Benakis et al. showed that intestinal dysbiosis affects Treg and IL-17 γδT cells and found that intestinal lymphocyte migration to the ischemic brain may be associated with increased infarct volume, while antibiotic-induced changes in the intestinal microbiota significantly reduced ischemic brain injury in mice (Benakis et al., 2016). Singh et al. observed dysbiosis of the gut microbiota by pressure-mediated intestinal palsy after CI, which in turn was causally associated with changes in T cell homeostasis, induction of the pro-inflammatory response, and worsening of stroke outcomes. Fecal microbiota transplantation (FMT) will contribute to improved stroke outcomes (Singh et al., 2016).

In addition, gut-derived damage-associated molecular patterns (DAMPs) and cytokine storms may affect the outcome of CI by regulating CNS antigen specific immune response. Tascilar et al. found that some intestinal bacteria in a mouse model of occlusion of the middle cerebral artery were translocated from the gut to the blood prior to the onset of the symptoms of CI. In addition, intestinal bacteria were transferred from the gut to the testes and other organs after CI in mice, and systemic inflammatory reactions occurred in some models later, causing post-stroke infection. However, the exact mechanism involved in this process remains unclear and needs further study (Tascilar et al., 2010). The gut microbiota of young mice with CI was altered, and in aged mice, the ratio of the two main bacterial phylums Firmicutes to Bacteroides phylum (F:B) increased approximately 9-fold compared to young mice, indicating dysbiosis. After altering the microbiota of young mice by fecal transplant gavage to increase their F:B ratio by about 6 times, cytokine levels were found to be significantly increased (Spychala et al., 2018). As a result, intestinal dysbiosis can create a vicious pro-inflammatory cycle that reduces the prognosis after stroke.

Regulation of the gut microbiota by antibiotics is a proven strategy, which can remove or prevent bacterial colonization in the human body, greatly affect the composition of the gut microbiota and reduce its biodiversity (Angelucci et al., 2019). The feasibility of alleviating CI by regulating intestinal microbial imbalance through antibiotics is being verified by various studies. Benakis et al. treated male mice with either a combination of antibiotics or a single antibiotic, respectively, and found that mice treated with a combination of antibiotics showed a significant reduction in infarct volume during the acute phase of stroke, while a single antibiotic treatment with ampicillin or vancomycin also reduced infarct volume and improved motor sensory function within 3 days after stroke (Benakis et al., 2020). Chen et al. applied antibiotics to CI rats, which decreased the α diversity of the intestinal microbiome, infarcted volume, and significantly increased acetic acid and valeric acid levels in ischemic rats (Chen et al., 2019). However, to date, there is no clear evidence to support prophylactic antibiotic therapy in the first hours after ACI to control ecological dissonance (Ulm et al., 2017). Two large randomized controlled phase III clinical trials did not show an improvement in results after ACI with prophylactic administration, and according to the results, antibiotics are not recommended for the prevention of CI (Kalra et al., 2015; Westendorp et al., 2015).

FMT is defined as the transfer of healthy gut bacteria via donor stool to a patient, aiming to obtain therapeutic benefits by directly altering or normalizing the gut microbiota of recipients (Khoruts and Sadowsky, 2016). Recent studies have confirmed FMT as a possible strategy to regulated intestinal ecological disorders in patients with neuropsychiatric disorders and patients affected by CI (Evrensel and Ceylan, 2016). Wang et al. performed FMT in a CI mouse model to verify the influence of different sexes microbiomes on the prognosis of CI. The results showed increased survival, reduced infarct size, improved performance in behavioral tests, increased release of beneficial metabolites, and reduced levels of inflammation in mice receiving the female gut microbiome. In contrast, mice that received the male microbiome were less effective in preventing brain damage and restoring neural function (Wang et al., 2022c). Chen et al. showed that FMT intervention significantly changed the intestinal microbiome composition of CI, reduced pathogenic bacteria and increased beneficial bacteria, and thus reduced neurological function damage, eliminated cerebral edema and reduced infarct volume. Further studies have shown that regulating SCFA levels such as isobutyric acid, butyric acid, and isovalerate may be the mechanism by which FMT alleviates CI (Chen et al., 2019).

The benefits of probiotics on host health have been extensively studied and clearly defined (Hill et al., 2014). Several studies have confirmed the efficacy and mechanism of probiotics in preventing CI by regulating the composition of the gut microbiota, improving intestinal barrier function, and regulating local and systemic immunity. Rodent models have verified the roles of Lactobacillus, Clostridium butyricum, and Bacillus licheniformis in CI (Fang et al., 2022). Through the regulation of TLR-4/NF-kappa B signaling, Lactobacillus reduced the extent of cerebral infarction, lowered oxidative stress, and blocked the death of brain cells, which improved neurobehavioral scores (Wanchao et al., 2018). After pretreatment with C. butyricum, butyrate content in the brain increases significantly, regulating CNS function and alleviating CI (Sun et al., 2016). “Synbiotics” is a combination of probiotics and matrices (Swanson et al., 2020). As opposed to inulin alone or SCFA-producing bacteria alone, Lee et al. discovered a synergistic impact of inulin and SCFA-producing bacteria, which improved neurodeficiency scores and behavioral outcomes in mice following stroke (Lee et al., 2020).

At the cellular level, excitotoxicity is a key cellular mechanism for cerebral ischemic injury (Rothman and Olney, 1986). This is triggered by metabolic homeostasis failure and secreted metabolites including glutamate, glycine, D-serine, and polyamines. Significant glutamate accumulation and down-regulation of glutamine ratios have been shown in ACI mouse models and in patients with cardiovascular diseases, including CI (Kimberly et al., 2013; Zheng et al., 2016).

Branched chain amino acids (BCAAs) are components of the glutamate/glutamine cycle between astrocytes and neurons and are essential for signaling in excitatory neurons (Salcedo et al., 2021). Metabolomic analysis of plasma in patients with mild and moderate ACI has found that reduced concentrations of BCAAs, including leucine, isoleucine, and valine, are associated with stroke severity and a worse prognosis (Kimberly et al., 2013). Wang et al. obtained the serum metabolic profile of stroke patients using the untargeted metabonomic method, and found that there were metabolic disorders in patients with ACI. Compared to HCs, the levels of 4-hydroxyproline, L-glutamine, L-arginine, and L-proline in patients with ACI were lower, revealing an increased risk of cerebrovascular diseases (Wang et al., 2020). Hu et al. directed injection liquid crystal mass spectrometry analysis of dried blood spots in patients with CI showed a significant increase in the ratio of tyrosine, citrulline, and proline, which also implied a decrease in neuronal autophagy, apoptosis, and platelet dysfunction (Hu et al., 2016).

As a subset of metabolomics, lipidomics shows functions similar to metabolomics (Lam et al., 2017). In recent years, the analysis of lipids has been greatly improved by matrix-assisted laser mass spectrometry and other techniques (Adibhatla et al., 2006). Several studies have shown that lipids play an important role in the occurrence and development of diseases. Lipids cross the blood-brain barrier more easily than proteins and enter brain cells, and the high abundance of polyunsaturated fatty acids in brain lipids makes the brain more susceptible to oxidative stress than most tissues and organs (Hamilton et al., 2007).

Animal models and human studies related to CI have shown that abnormal lipid metabolism is closely related to the prediction and prognosis of CI, including phospholipids, sphingolipids (SLs), and glycerides (Ye et al., 2022). Sheth et al. measured and evaluated the changes in plasma SL concentration in the mouse model of ACI, and the results showed that SLs were highly enriched in the brain, among which the most obvious changes were ceramide and sphingomyelin (SM), and the concentration of multiple SLs in the brain and plasma differed by more than 1000 times. The feasibility of this targeted lipid analysis was verified in patients with ACI (Sheth et al., 2015). Human serum metabolomics based on nuclear magnetic resonance was performed on blood phosphorus in patients with CI, and it was found that serum phosphatidylcholine, phosphoethanolamine and SM levels were lower in CI patients compared with healthy individuals (Grandizoli et al., 2014). Therefore, phosphorus-containing compounds in human serum can be considered important biomarkers in CI research.

Oxidative stress in cerebral ischemia is caused by the excessive production of oxygen derivatives and metabolic dysfunction. Stroke causes heterogeneous changes in tissue oxygenation and produces lactic acid as the end product, thus making the cytoplasmic environment acidic (Liu et al., 2004). Excess protons convert oxygen into hydrogen peroxide and reactive hydroxyl radicals (Shin et al., 2020). Jung et al. found that the levels of lactic acid, pyruvate, glycolic acid and formate in plasma and urine of patients with cerebral ischemia increased, while the levels of glutamine and methanol decreased, which reflected the oxidative stress state of cerebral ischemia (Jung et al., 2011).

In individuals with cerebral ischemia, oxidative stress and blood-brain barrier damage can also aggravate the inflammatory response (Sidorov et al., 2019). Glial cells, neutrophils, monocytes, and lymphocytes are seen in higher concentrations with the activation of inflammation, along with pro-inflammatory cytokines and metabolites that are connected to inflammation (Shin et al., 2020). The increase of C4/C2 could indicate the deficiency of short-chain acyl-coA dehydrogenase. This is closely related to the inactivation of peroxisome proliferator-activated receptor alpha, which may reflect platelet dysfunction. The increase of alanine level is believed to be related to the increase of carnosin content in blood samples of cerebral ischemia patients, and is closely related to neuronal autophagy (Hu et al., 2016).