Under normal oxygen levels, the proline residue of the HIF-1α protein binds to Von Hippel-Lindau proteins in the presence of oxygen, iron (II) and α-ketoglutarate, leading to its rapid degradation through the ubiquitin-proteasome pathway (14). By contrast, hypoxic conditions prevent the hydroxylation of HIF-1α, allowing it to accumulate, translocate to the nucleus and bind with HIF-1β, thereby triggering the transcription of genes related to hypoxia, such as VEGF (15). The brain is highly sensitive to oxygen and nutrient fluctuations, requiring efficient blood supply. To support its metabolic needs, the glucose transporter type 1 (GLUT1) protein is expressed in ECs and neurons, facilitating glucose transport and sustaining glycolytic processes. Following IS, neuronal cells in the ischemic penumbra have an increased demand for nutrients, which cannot be transported without ECs. HIF-1α activation can maintain redox homeostasis by facilitating glucose transport and glycolysis. Studies have revealed that hypoxic preconditioning helps reduce cortical neuronal loss in rats with traumatic brain injury (16,17). This protective effect is primarily linked to the upregulation of HIF-1α, which in turn stimulates the expression of GLUT1 and 3, enhancing glucose uptake by neurons. The phosphatidylinositol 3-kinase/protein kinase B/mammalian target of rapamycin (PI3K/AKT/mTOR) signaling pathway plays a crucial role in vascularization, including the survival, migration and angiogenesis of ECs. That activation of the PI3K/AKT/mTOR pathway further stimulates its downstream target, HIF-1α, which regulates the expression of VEGF, a key factor that promotes vascularization following IS (18). In addition, receptors such as the epidermal growth factor receptor, fibroblast growth factor receptors and ΙL-6 receptor exert neuroprotective effects against cerebral ischemia by activating the PI3K/AKT/mTOR pathway, leading to increased levels of HIF-1α (19). HIF-1α, in turn, promotes angiogenesis in ischemic tissues by increasing VEGF expression (20). In ECs, the PI3K/AKT pathway is also involved in angiogenesis through its regulation of nitric oxide (NO) signaling, which is modulated by endothelial NO synthase (eNOS). Evidence suggests that NO donors can enhance the transcriptional activity and expression of HIF-1α, thereby increasing VEGF mRNA levels (21). These findings indicated that HIF-1α plays a pivotal role in regulating EC dysfunction under oxidative stress following IS, largely through stimulating angiogenesis. The mechanism of HIF-1α and ECs angiogenesis-related factors is demonstrated in Fig. 2.

Pro-inflammatory molecules, such as TNF-α, IL-1 and IL-6, are key contributors to the development of cerebral infarction. Although these factors exacerbate tissue damage mainly by promoting inflammatory responses, they can, in some cases, participate in angiogenesis by modulating angiogenesis-related signaling pathways.

TNF-α is a cytokine that exerts numerous pro-inflammatory effects and is pivotal in both pathological and physiological processes (22). Following IS, TNF-α is activated within 1 h, reaches its peak between 6 and 10 h, and typically diminishes within 1 to 2 days (23). Increased TNF-α exerts both neuroprotective and neurotoxic effects following IS (23). TNF-α exerts its effects by binding to two receptors, TNF receptor 1 (TNFR1) and TNFR2, both of which are involved in hind limb ischemia-induced angiogenesis (24). When TNF-α interacts with TNFR1, it triggers the expression of pro-erythropoietin (EPO) in cerebral ECs by increasing the EPO receptor (EPOR) levels. EPO then promotes angiogenesis by upregulating EPOR, which enhances the activation of signaling pathways such as VEGF/VEGFR2 and angiopoietin-1/Tie2 (Ang1/Tie2) (25). Furthermore, TNF-α directly influences ECs to regulate angiogenesis via VEGF signaling activation. In addition, TNF activates several signaling pathways, including NF-κB and AKT. NF-κB activation by TNF can trigger AKT signaling, and both the NF-κB and PI3K/AKT pathways are crucial for the upregulation of TNF/TNFR1 in EPOR (26). These findings suggested that TNF-α may play a neuroprotective role by promoting angiogenesis through EPO and its associated signaling mechanisms.

IL-1. The pro-inflammatory cytokine IL-1 plays a central role in cerebrovascular inflammation following ischemic injury. After cerebral ischemia, microglia, astrocytes and ECs release two isoforms of IL-1, IL-1α and IL-1β, which primarily target ECs and astrocytes (27,28). IL-1 stimulates the expression of LG3, a C-terminal fragment of perlecan, in brain cell cultures. Perlecan is a key component of the basement membrane of ECs in mature tissues, and LG3 has been identified as a potentially neuroprotective and pro-angiogenic fragment of the extracellular matrix (29). LG3 can mitigate β-amyloid-induced neuronal toxicity by binding to the α2β1 integrin (30). In addition, IL-1 promotes the expression of pentraxin-3 (PTX3), an acute-phase protein involved in brain repair processes following ischemia. PTX3 is crucial for mechanisms such as neurogenesis and angiogenesis in the post-ischemic brain (31). After 14 days of MCAO, VEGF exerts a potent pro-angiogenic effect through the activation of VEGFR2, which is upregulated after ischemia and reduced in PTX3 knockout mice (32).

The pro-inflammatory actions of IL-1β are mediated via its receptor, IL-1R1. The inhibition of IL-1β binding to IL-1R1 can prevent cerebral edema and brain tissue damage in experimental IS models. IL-1β stimulates the chemokines SDF-1/CXCL-12, which accumulates at the BBB, promoting leukocyte infiltration into the CNS, along with microvascular leakage and cerebral edema (33). Studies on the impact of IL-1β on trans-endothelial electrical resistance in human cerebral microvasculature have revealed that IL-1β disrupts the endothelial barrier through the PKC-dependent phosphorylation of PKC-θ and ZO-1(34). As IL-1β is a key inflammatory mediator, targeting the inhibition of PKC-θ or ZO-1 phosphorylation offers a potential strategy to prevent BBB disruption during neuroinflammation (35). In addition, macrophage-derived IL-1β enhances the expression of the pro-angiogenic isoform VEGF-A165a, while suppressing the anti-angiogenic VEGF-A165b through the activation of signal transducer and activator of transcription-3 (STAT-3) and NF-κB pathways. This shift promotes VEGF expression and contributes to inflammatory vascularization (36).

IL-1α has been revealed to be significantly more potent than IL-1β in activating ECs, as evidenced by the increased expression of the pro-angiogenic chemokine CXCL-1. In addition, IL-1α stimulates a strong, concentration-dependent expression of the angiogenic mediator IL-6(37). It also promotes EC proliferation, migration and the formation of tubular structures. These responses can be blocked by the IL-1 receptor antagonist. Thus, while IL-1-driven inflammation can exacerbate ischemic injury, it also plays a dual role by enhancing the brain repair mechanisms and supporting functional recovery.

IL-6. IL-6 is a multifunctional cytokine and an important messenger molecule between leukocytes, vascular ECs and thin-walled tissue-resident cells, produced by several types of cells, such as monocytes, macrophages, adipocytes, hematopoietic cells and ECs (38). The function of IL-6 is mediated through a unique receptor system consisting of two functional proteins: An IL-6-specific receptor (IL-6R) and a second glycoprotein, gp130. Signaling through gp130 is mediated by two pathways: The JAK-STAT and the Ras-MAPK pathway (39). In the vasculature, IL-6 can act directly or indirectly on vascular ECs. IL-6 preconditioning induces VEGF secretion from neural stem cells via STAT3, which promotes angiogenesis in the ischemic brain (40). A related study (41) found that two days after mild transient cerebral ischemia, wild-type mice exhibited a significant early increase in IL-6 gene transcription and protein levels in the ischemic brain. This was accompanied by an early upregulation of mRNAs for angiogenesis-related genes, such as VEGFR2 and eNOS. In mice with MCAO, the increase in circulating VEGF was diminished following IL-6 knockdown at 48 h (42). Monocyte-derived IL-6 plays a critical role in programming microglia to repair the damaged brain vasculature. Cerebrovascular repair was absent in IL-6 knockout mice or those lacking microglial IL-6Ra expression, but could be restored through exogenous IL-6 administration (43).

The aforementioned studies suggested that IL-6 promotes angiogenesis following cerebral infarction, thereby providing long-term histological and functional protection. ECs and inflammatory cytokines are shown in Fig. 3.

The EC membrane serves as the interface between ECs and the external environment, facilitating material exchange and signal transmission. The integrity and dynamic changes of the EC membrane significantly influence the activity and role of different adhesion molecules. These molecules, expressed on the ECs membrane, are critical in vascular biology as they regulate blood flow, immune cell migration and inflammatory responses. Specifically, adhesion molecules are vital for leukocyte infiltration into the CNS. The molecular interactions between ECs and circulating leukocytes are primarily mediated by three categories of adhesion molecules: Selectins such as E-, P- and L-selectins, immunoglobulin superfamily members such as intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) and integrins.

Selectins are membrane-bound glycoproteins with three main members: E-selectin (CD62E), P-selectin (CD62P) and L-selectin (CD62L) (44). The molecular mechanisms of leukocyte infiltration into ischemic tissues involve these selectins. CD62E, also known as E-selectin, plays a crucial role in the adhesion of leukocytes to ECs, and its involvement in the homing and angiogenesis of EPCs has been experimentally studied. CD62E is synthesized in response to inflammatory stimuli, such as TNF-α and IL-1, and is expressed on the EC membrane within hours of stimulation. CD62P or P-selectin, is found on ECs and platelet granule membranes, and is rapidly expressed on the outer cell membrane upon activation by factors such as thrombin or histamine. CD62L is present on lymphocytes, neutrophils and monocytes, and following cell activation it is cleaved from the membrane through proteolytic processes.

Immunoglobulin superfamily. The immunoglobulin superfamily consists of a group of cell surface and soluble proteins, several of which serve as adhesion molecules. A total of 5 key members of this family include ICAM-1, ICAM-2, VCAM-1, platelet endothelial cell adhesion molecule 1 and mucosal vascular addressin in cell adhesion molecule 1. Among these, ICAM-1 and VCAM-1 are some of the most extensively studied adhesion molecules in the context of IS. The expression of these EC adhesion molecules is a crucial step in the recruitment of circulating leukocytes to areas of inflammation. Hypoxic conditions and cytokines, such as IL-1β and TNF-α, significantly enhance the expression of these molecules (45).

The role of ICAM-1 in regulating cerebral leukocyte recruitment during neuroinflammation and ischemia has been clearly established through studies involving ICAM-1 knockout mice and the use of ICAM-1 blocking antibodies (46). In vivo research in mice has revealed that TNF-α triggers the upregulation of both ICAM-1 and VCAM-1. The expression levels of these molecules begin to rise between 2 and 5 h after brain injury, peak between 5 and 9 h, and remain elevated above baseline levels for at least 24 h (47). Of note, the increases in ICAM-1 and VCAM-1 expression are dose-dependent, with a significant upregulation at 5 µg/kg and a maximal increase observed at doses between 10-25 µg/kg.

Correlative studies have revealed that VEGF increases VCAM-1 and ICAM-1 protein levels and promotes leukocyte adhesion in an NF-κB-dependent manner (48). ICAM-1 expression influences VEGF-A-induced eNOS activity and angiogenesis by regulating endothelial glutathione levels (49). Studies also have revealed that both VCAM-1 and integrin α4 (ITGA4) are upregulated by TNF-α in ECs. sVCAM-1-induced angiogenesis is facilitated by the upregulation of VEGF through the p38 MAPK/FAK signaling pathway. The sVCAM-1/ITGA4 pathway may play a role in inflammatory angiogenesis (50).

The aforementioned findings suggested that inflammatory cytokines promote the increase in the expression of ICAM-1 and VCAM-1, which enhances the interaction between ECs and circulating leukocytes, thereby promoting angiogenesis following IS.

Integrins. Integrins are heterodimeric proteins made up of distinct α-subunits paired with common β-subunits. The β-subunits are categorized into three subfamilies: β1, β2 and β3 integrins. Integrins in the β1 subfamily play a key role in connecting to the extracellular matrix, binding to collagen, laminin and fibronectin. By contrast, β2 integrins are crucial for mediating the adhesion between leukocytes and ECs. β3 integrins, also referred to as integrin αvβ3 or cytokines, are involved in blood clot formation and stabilization. These integrins are vital for regulating processes such as hematopoiesis, leukocyte recruitment and inflammatory responses (51).

Brain ECs demonstrate minimal expression of adhesion molecules, which prevents peripheral immune cells from crossing into the CNS (25). β2 Integrin, a key integrin subunit, plays a significant role in directing EPCs to areas of active vascular angiogenesis. Known as a leukocyte-specific receptor, β2 integrin interacts with various ICAM family members and polysaccharides (52).

The impact of neuroinflammatory mechanisms (whether detrimental or protective) depends on the timing following cerebral ischemia. Inflammation may exacerbate ischemic injury in the early stages of IS, whereas the inflammatory response may protect neurons by promoting neurogenesis, angiogenesis and neuroplasticity in the later stages. Integrins such as α5β1 and αvβ3, play a crucial role in regulating the processes of angiogenesis and inflammation following cerebral ischemia (53). The Ang1/Tie2 signaling pathway and the integrin α5β1 interact with ECs following IS, promoting angiogenesis (54). VEGF upregulation by α5β1 and αvβ3, and their ligand-endo-ligand proteins in the ischemic penumbra, stimulate EC proliferation. TNF-α or FGF-2-induced angiogenesis is dependent on αvβ3, while VEGF and TGF-α-induced angiogenesis is dependent on αvβ5, suggesting that αv integrins play a key role in potent angiogenesis (55).

Adhesion molecules on ECs play a crucial role in maintaining vascular health and function. Targeting these molecules to modulate their expression can reduce inflammation and improve EC function. For example, the development of small-molecule inhibitors targeting ICAM-1 or VCAM-1 may help suppress intravascular inflammation and reduce cerebrovascular lesions (56). The development of therapies that promote EC repair and regeneration may help to improve the function of the vascular endothelium and provide a new direction in the treatment of cerebrovascular diseases. For example, stem cell therapy (57) or gene therapy (58) is used to promote endothelial repair and restore the normal function of blood vessels. The association between ECs and adhesion factors is demonstrated in Fig. 4.