Canonical NF-κB signaling pathway and GRO-α/CXCR2 axis are activated in unruptured intracranial aneurysm patients

Abstract

Activation of the nuclear factor kappa-B (NF-κB) stimulates the production of pro-inflammatory molecules involved in the formation of intracranial aneurysms (IA). The study aimed to assess the NF-κB p65 subunit and the GRO-α chemokine and its receptor CXCR2 concentrations in unruptured intracranial aneurysm patients (UIA, n = 25) compared to individuals without vascular changes in the brain (n = 10). It was also analyzed whether tested proteins are related to the size and number of aneurysms. Cerebrospinal fluid (CSF) and serum protein levels were measured using the ELISA method. Median CSF and serum NF-κB p65 concentrations were significantly lower, while median CSF GRO-α and CXCR2 concentrations were significantly higher in UIA patients compared to the control group. CSF and serum NF-κB p65 concentrations negatively correlated with the number of aneurysms. In UIA patients the median GRO-α concentration was two-fold and CXCR2 almost four-fold higher in CSF compared to the serum value. CSF GRO-α concentration positively correlated with the size of aneurysms.Significantly decreased CSF NF-κB p65 and significantly increased CSF GRO-α and its CXCR2 receptor concentrations in UIA patients compared to the control group may altogether suggest that the canonical NF-κB signaling pathway is activated and its target pro-inflammatory genes are highly expressed in UIA patients. However, to unequivocally assess the involvement of the classical NF-κB pathway with the participation of the NF-κB p65 subunit and the GRO-α/CXCR2 axis in the formation of IA, further in vivo model studies are needed.

Figure 1

(A–H) Cerebrospinal fluid, serum, and Quotient results for NF-κB p65 in unruptured intracranial aneurysm patients and the control group. The Quotient was calculated by dividing the CSF protein value by the serum protein value. Statistical significance: * p < 0.05, ** p ≤ 0.01, **** p ≤ 0.001. CSF, Cerebrospinal fluid; NF-κB p65, nuclear factor kappa-B p65 subunit; UIA, unruptured intracranial aneurysm.

Figure 2

(A–E) Cerebrospinal fluid, serum, and Quotient results for GRO-α in unruptured intracranial aneurysm patients and the control group. The Quotient was calculated by dividing the CSF protein value by the serum protein value. Statistical significance: * p < 0.05, **** p ≤ 0.001. CSF, Cerebrospinal fluid; GRO-α, GRO alpha chemokine; NF-κB p65, nuclear factor kappa-B p65 subunit; UIA, unruptured intracranial aneurysm.

Figure 3

(A–E) Cerebrospinal fluid, serum, and Quotient results for CXCR2 in unruptured intracranial aneurysm patients and the control group. The Quotient was calculated by dividing the CSF protein value by the serum protein value. Statistical significance: * p < 0.05, **** p ≤ 0.001. CSF, Cerebrospinal fluid; CXCR2, C-X-C Motif Chemokine Receptor 2; UIA, unruptured intracranial aneurysm.

Figure 4

Areas under the ROC curves (AUCs) for NF-κB p65, GRO-α, and CXCR2 Quotients in differentiating unruptured intracranial aneurysm patients from individuals without vascular lesions in the brain. Quotients were calculated by dividing the CSF protein value by the serum protein value. For NF-κB p65 Quotient the AUC = 0.772, cut-off = 0.37; for GRO-α Quotient the AUC = 0.728, cut-off = 1.60; for CXCR2 Quotient the AUC = 0.732, cut-off = 2.07.

Figure 5

A schematic presentation of the possible role of the NF-κB p65 subunit and the GRO-α/CXCR2 axis in the formation of IA. The first step initiating the formation of IA is the activation of the canonical/classical NF-κB pathway in the arterial bifurcationas as the result of hemodynamic stress. NF-κB activation occurs through the phosphorylation and proteasomal degradation of IκB. An active form of NF-κB p50/p65 heterodimer bind to DNA to function as a transcription factor of pro-inflammatory cytokine genes, including chemokine GRO-α. Secretory chemokine GRO-α, via its CXCR2 receptor, acts as a chemoattractant for leukocytes and increases endothelial cell proliferation, which initiates the formation of IA. As a result of forming IA, also microglia and astrocytes could secrete GRO-α, which in turn enhances leukocyte trafficking and endothelial cell proliferation. CXCR2, C-X-C Motif Chemokine Receptor 2; GRO-α, GRO alpha chemokine/C-X-C motif ligand 1 (CXCL1); IA, intracranial aneurysms; IκB, IkappaB kinase; NF-κB p50, nuclear factor kappa-B p50 subunit; NF-κB p65, nuclear factor kappa-B p65 subunit; NF-κB, nuclear factor kappa-B.