Currently, excitotoxicity, calcium dysregulation, oxidative and nitrosative stress, cortical spreading depolarizations, and inflammation are used to explain the pathophysiology of IS [19, 20]. Ischemic neuroinflammation is a pathological process of IS that significantly affects clinical prognosis [21]. Notably, cytokines, secreted by different brain immune cells, are key players in the inflammatory response, exhibiting both proinflammatory and anti-inflammatory effects [22, 23]. Fig. 1 shows the inflammatory response of immune cells.

Fig. 1.

Pathophysiological process of neuroinflammatory response after ischemic stroke. After ischemic brain injury, DAMPs are released during cell ischemia, hypoxia, and necrosis, promoting the activation of microglia, astrocytes, brain endothelial cells, and perivascular immune cells. Microglia polarize into classical (M1) or alternative (M2) microglia and secrete proinflammatory cytokines, anti-inflammatory cytokines, adhesion molecules, and chemokines. Activated astrocytes initially secrete proinflammatory cytokines, followed by anti-inflammatory cytokines, chemokines, and adhesion molecules. Proinflammatory cytokines contribute to the BBB breakdown and exacerbate the inflammatory response, while anti-inflammatory cytokines counteract this process and play a neuroprotective role. Adhesion molecules and chemokines facilitate the infiltration of peripheral immune cells into the BBB and promote the inflammatory response. DAMPs, damage-associated molecular patterns; BBB, blood-brain barrier; TLRs, Toll-like receptors; M1, classical microglia; M2, alternative microglia.

After ischemic brain injury, ischemia-anoxic necrosis cells release a series of danger signals commonly called damage-associated molecular patterns (DAMPs), such as the high-mobility group box 1 protein (HMGB1) [24, 25, 26]. These DAMPs are released into the extracellular environment to activate the innate immune system and further stimulate the inflammatory cascade reaction [27]. Toll-like receptors (TLRs), located on microglia, astrocytes, perivascular macrophages, and brain endothelial cells, detect DAMPs and activate corresponding cells in response [28, 29, 30, 31].

Microglia, the brain’s most predominant resident macrophages, account for approximately 10% of all brain tissue cells that become highly activated early after stroke [32, 33, 34]. After activation, microglia polarize into two types, classical (M1) and alternative (M2) microglia, which play different roles, secreting cytokines that promote or relieve inflammation and affecting tissue repair [35, 36]. This process increases microglial phagocytosis activity [37]. Currently, microglial cells secrete a variety of inflammatory cytokines and chemokines, including interleukin(IL)-1$\beta$, IL-4, IL-6, IL-8, IL-10, IL-12, IL-15, tumor necrosis factor (TNF)-$\alpha$, transforming growth factor (TGF)-$\beta$, macrophage inflammatory protein (MIP)-1, monocyte chemoattractant protein-1 (MCP-1), chemokines interferon-$\gamma$-inducible protein 10 (CXCL10, also called IP-10), and proteolytic enzymes including matrix metalloproteinase (MMP)-3 and MMP-9 [38, 39, 40].

In the central nervous system, astrocytes are the main type of glial cells, accounting for approximately 50% of brain cells. In addition to playing a supporting role, astrocytes are important regulatory factors of brain function [41, 42, 43]. When activated by proinflammatory factors secreted by activated microglia, astrocytes lose their neuroprotective and homeostatic ability and induce neuron and oligodendrocyte death [44]. After brain tissue ischemia, astrocytes reduce brain nerve damage by producing antioxidants and glutamine and regulating extracellular potassium concentration [45]. Activated astrocytes secrete large amounts of inflammatory cytokines and chemokines, such as IL-1, TNF-$\alpha$, interferon-$\gamma$ (IFN-$\gamma$), and MCP-1, which contribute to BBB destruction [46, 47]. While astrocyte death leads to decreased expression of tight junction proteins, leading to BBB breakdown and brain tissue edema [48]. Inflammatory factors such as IL-1, TNF-$\alpha$, and MCP-1 increase in the early stages of cerebral ischemic injury, while IL-6 and macrophage migration inhibitory factors (MIF) increase at later stages [49]. Moreover, proinflammatory factors enhance the expression of adhesion molecules, including E-selectin, P-selectin, intercellular cell adhesion molecule (ICAM)-1, ICAM-2, and vascular endothelial cell adhesion molecule (VCAM)-1 [50, 51, 52, 53].

Neutrophils are one of the peripheral immune cells. Neutrophils also release cytokines that promote glial scarring, phagocytic debris, and edema resolution [54]. Following BBB damage, neutrophils penetrate the brain through the action of cell adhesion molecules, including selectin, integrin, and immunoglobulin [55]. Neutrophils have a strong ability to destroy brain tissue, as they generate reactive oxygen species (ROS) and proteolytic enzymes, inducing direct neurotoxic effects. Additionally, their accumulation indirectly leads to cerebral ischemia and hypoxia by blocking capillary blood flow [56, 57].

Another peripheral cells, (such as mononuclear phagocytes, natural killer cells (NK cells), and lymphocytes) also release cytokines and participate in ischemic inflammation [23, 58]. Monocytes are innate immune cells that serve as important effectors and regulatory factors in inflammation [59], crossing the BBB, accumulating in damaged brain tissue, and exhibiting microglia-like phenotypic changes [60, 61]. NK cells are important innate immune cells in peripheral blood that mobilize quickly in the initial stage of inflammatory responses, thereby promoting neuroinflammatory and inducing neuronal death [62].

A growing body of literature has begun to elucidate that lymphocytes can assume in IS pathology. Lymphocytes cross the BBB and enter ischemic brain parenchyma under the influence of P-selectin, VCAM-1, and ICAM-1 signaling [63]. The degree of lymphocyte infiltration depends on changes in BBB permeability [64]. Immune system activation mediated by lymphocyte infiltration can have both harmful antigen-specific autoreactive responses and cellular protection in IS [65]. During this process, helper T cells are divided into Th1 cells secreting proinflammatory cytokines (such as IFN-$\gamma$, TNF-$\alpha$, and lymphotoxin alpha (LT-$\alpha$)), Th2 cells secreting anti-inflammatory cytokines (such as IL-4 and IL-10), and TH17 cells secreting IL-17, IL-21, and IL-22 cytokines [66]. $\gamma$$\delta$ T cells, associated with post-stroke neurotoxicity, are induced by macrophage-secreted IL-23 and DAMPs-activated surface TLRs [67]. Activated $\gamma$$\delta$ T cells migrate to the ischemic tissue and produce IFN-$\gamma$ and IL-17 [68]. IFN-$\gamma$, mainly secreted by T cells, mediates delayed neurotoxic effects in IS [69, 70]. Regulatory T cells (Treg cells) exhibit stronger anti-apoptotic properties compared to other cells [71] and secrete cytokines such as IL-10, which have protective effects after cerebral ischemia. Treg cells reduce the early invasion and activation of neutrophils and T cells. When Treg cells are depleted, the expressions of TNF-$\alpha$, IL-1$\beta$, IFN-$\gamma$, and other proinflammatory cytokines increase in the brain [72]. Regulatory B cells also have beneficial effects on the ischemic brain by secreting IL-10 [73]. Studies have shown that lack of B cells increases migration cross the BBB and decreases IL-10 production. Conversely, infiltration of B cells into the brain reduces brain infarct size and peripheral T cell activation [73].

Brain ischemia triggers a surge in inflammatory cytokine production by various cell types, including microglia, astrocytes, endothelial cells, and neurons, with contrasting outcomes. Proinflammatory cytokines (such as TNF-$\alpha$ and IL-1) exacerbate stroke-related brain injury, while anti-inflammatory cytokines (such as IL-10 and TGF-$\beta$) alleviate cerebral ischemia injury [23, 55, 74]. Table 1 (Ref. [8, 22, 55, 69, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, 86, 87, 88, 89, 90, 91, 92, 93, 94, 95, 96, 97, 98, 99, 100, 101, 102, 103, 104, 105, 106, 107, 108, 109, 110, 111, 112, 113, 114, 115, 116, 117, 118, 119, 120, 121, 122, 123, 124]) outlines the role of inflammatory mediators.

Previous studies have associated differences in the production of inflammatory to infarct size following cerebral ischemia. Proinflammatory factors can increase cell edema and death, while the expression of anti-inflammatory factors can partially alleviate nerve damage [125, 126]. IL-1 is widely produced in nerve cells, microglia, and astrocytes after brain tissue ischemia. It is an important proinflammatory cytokine in the inflammatory response [55]. IL-1 can promote neuronal death by inducing calcium ions to enter cells by acting on N-Methyl-D-aspartic acid receptors (NMDARs) [78]. IL-1 also activates phospholipase A2, disrupting cell phospholipase bilayers, inducing apoptosis, and promoting the release of metabolites such as prostaglandins and leukotrienes, which can induce BBB breakdown and promote inflammatory response [93, 127]. Furthermore, IL-1 induces BBB breakdown and promotes leukocyte infiltration through adhesion molecules on endothelial cells [55, 128, 129, 130]. Adhesion molecules act as ligands involved in leukocyte chemotaxis and inflammatory responses by binding to leukocyte integrin receptors attached to endothelial cells [20]. Moreover, inflammatory stimulation results in a significant upregulation of ICAM-1 and animal studies have shown that artificial intervention with IL-1 receptor antagonists can reduce cerebral infarct size [131, 132, 133].

IL-6 is a proinflammatory cytokine that is highly expressed after a stroke event [134] and is associated with brain tissue necrosis and poor clinical prognosis [55, 94, 135]. However, there is another view that IL-6 has an anti-inflammatory effect by promoting tissue recovery and reducing infarct severity [136, 137, 138]. Currently, the known anti-inflammatory action mechanisms include inhibiting inflammatory factors, inhibiting TNF-$\alpha$ expression, inducing the production of IL-1 receptor antagonists, and inducing neutrophil apoptosis [95, 139]. Therefore, the role of IL-6 in the neuroinflammatory pathophysiological of IS is complex.

TNF-$\alpha$ is an important proinflammatory cytokine produced by microglia, monocytes, and other immune cells during acute inflammation, resulting in cell necrosis or apoptosis through intracellular signaling pathways [140]. TNF-$\alpha$ level was positively correlated with cerebral infarction volume [141, 142]. Artificial use of anti-TNF-$\alpha$ and TNF-$\alpha$ binding protein could decrease tissue necrosis and infarct size [143, 144, 145, 146]. TNF-$\alpha$ can activate microglia and astrocytes, induce the expression of adhesion molecules in brain endothelial cells, destroy the BBB, promote leukocyte infiltration, and affect glutamate transfer and synaptic plasticity [22, 75].

IL-10 is an anti-inflammatory cytokine widely present in astrocytes, mononuclear macrophages, Th cells, and B cells, and animal experiments have demonstrated that mice with IL-10 overexpression have smaller infarct size and limited apoptosis [97]. Previous research showed that IL-10 can reduce infarct size after focal ischemia, reduce leukocyte and monocyte/macrophage infiltration, and mitigate nerve damage caused by inflammatory cytokines [72, 147, 148].

TGF-$\beta$, a neuroprotective factor, promotes tissue regeneration and neurological recovery [55, 149, 150]. Experimental results showed that TGF-$\beta$ can promote endothelial cell angiogenesis, stimulate M2 polarization of microglia cells, and inhibit neuron damage [151]. Moreover, TGF-$\beta$ stabilizes the BBB during early stroke [152, 153].

Nuclear factor kappa-B (NF-$\kappa$B), a heterogeneous transcription factor activated after cerebral ischemia, induces inflammation [154, 155, 156]. Studies have shown that NF-$\kappa$B influences microglia activation and polarization [157]while promoting the expression of inflammatory cytokines, MMPs, and adhesion molecules in inflammatory response [158, 159, 160]. Multiple pathways, including the suppressor of cytokine signaling 1/janus kinase/signal transducer and activator of transcription (SOCS-1/Janus/STAT) signaling pathway, are involved [161].

MMPs are a family of proteolytic enzymes which can induce BBB breakdown [162]. During the acute phase of IS, MMP expression increases [163, 164]. MMPs are found in microglia, astrocytes, and endothelial cells. Moreover, MMPs activate microglia, macrophages, and neutrophils, further promoting MMP secretion and leading to vascular and BBB damage [165]. Experimental studies have shown that silencing MMP-Ⅸ and MMP-Ⅲ gene expression significantly alleviated BBB damage and edema [166, 167].

CircRNAs are non-coding RNA molecules that form covalent ring structures without 5${}^{\prime }$ terminal cap or 3${}^{\prime }$ terminal poly (A) tails [168]. CircRNAs were first discovered in plant viroids in 1976 and were widespread in the cytoplasm of eukaryotes [169, 170, 171, 172, 173, 174, 175]. Initially considered as by-products of mis-splicing [176], circRNAs currently have various functions (Fig. 2), including (i) acting as microRNA (miRNA) sponges: circRNAs contain miRNA binding sites, competitively binding miRNAs to indirectly regulate their transcription and translation [177]; (ii) impacting RNA polymerase II transcription: circRNAs can form R-loops with their producing locus or coactivate transcription factors (TFs) to modulate transcription [178, 179]; (iii) regulating splicing: circRNA back-splicing sites can compete with pre-mRNA splicing, influencing mRNA synthesis [180, 181]; (iv) absorbing or combining proteins: circRNAs containing protein-binding sites can bind to proteins, influencing their participation in other neuronal functions [180, 182, 183, 184]; (v) forming functional circRNA-protein (circRNP) complexes: several circRNAs can form circRNP complexes with RNA binding proteins (RBPs), thereby modulating signaling pathways [185, 186, 187]; (vi) affecting mRNA expression: partial circRNAs can impact mRNAs’ stability and translation by directly binding to mRNAs [188, 189]; and (vii) protein translation: some circRNAs through splicing-dependent and cap-independent mechanisms, can be translated into proteins [190, 191, 192].

Fig. 2.

Biogenesis and function of circRNAs. (A) Lariat-driven circularization model: the 3${}^{\prime }$ splice donor of exon 1 and the 5${}^{\prime }$ splice acceptor of exon 4 links up end-to-end by exon skipping and form an exon-containing lariat structure. Finally, the ecircRNA forms after intron removal. (B) Intron pairing-driven circularization model: direct base pairing of introns forms a circulation structure, thereby forming ecircRNA or EIciRNA after intron removal. (C) RBP-dependent cyclization model: RBPs bridge two flanking introns close together and then remove introns to form circRNAs. (D) ciRNA formation model: the elements near the splice site escape debranching stably to form the intron lariat from the splicing reaction. (E) tRNA intronic circRNA: tricRNAs derived from introns removed during precursor-tRNA (pre-tRNA) splicing. (F) Act as microRNA (miRNA) sponges. (G) Impact RNA Pol II transcription. (H) Regulate splicing. (I) Absorb or combine proteins. (J) Form functional circRNP complexes. (K) Interact with mRNAs to affect their expression. circRNAs, Circular RNAs; ecircRNAs, exonic circRNAs; EIciRNAs, exon-intron circRNAs; RBP, RNA binding protein; ciRNAs, circular intronic RNAs; tricRNAs, tRNA intronic circular RNAs.

CircRNAs can be categorized into four groups based on their biogenesis: exonic circRNAs (ecircRNAs), synthesized from only the exon sequence, circular intronic RNAs (ciRNAs) containing introns, exon-intron circRNAs (EIciRNAs) containing both exon and intron sequences, and tRNA intronic circular RNAs (tricRNAs) [10, 173, 193, 194, 195]. The biogenesis of circRNAs is illustrated in Fig. 2.