Fibroblast Growth Factor-18 (FGF-18) Attenuates ox-LDL-Induced Endothelial Inflammation and Monocyte Adhesion by Inhibiting TRIM28

Abstract

The adhesion of monocytes to vascular endothelial cells constitutes a fundamental early process in atherogenesis. Fibroblast growth factor-18 (FGF-18), known to signal through the fibroblast growth factor receptor 1 (FGFR1), has emerging roles in maintaining vascular homeostasis, but its precise function in endothelial inflammation remains unclear. The protective role of recombinant human FGF-18 (rhFGF-18) against oxidized low-density lipoprotein (ox-LDL)-induced endothelial injury and its mechanism were investigated. We found that ox-LDL downregulated phospho-FGFR1 in human aortic vascular endothelial cells (HAVECs) dose- and time-dependently. rhFGF-18 treatment markedly suppressed ox-LDL-induced upregulation of the scavenger receptor LOX-1 and key pro-inflammatory factors (TNF-α, MCP-1, COX-2, and PGE₂). Subsequently, rhFGF-18 reduced the expression of adhesion molecules VCAM-1 and ICAM-1, thereby decreasing THP-1 monocyte adhesion to HAVECs. Mechanistic investigations revealed that rhFGF-18 inhibits ox-LDL-induced phosphorylation and nuclear translocation of the transcriptional regulator TRIM28. Importantly, TRIM28 overexpression reversed the anti-inflammatory and anti-adhesive benefits of rhFGF-18. Collectively, this study identifies the FGF-18/TRIM28 axis as a crucial mechanism alleviating endothelial inflammation and monocyte adhesion, highlighting its potential as a therapeutic target for atherosclerosis.

Keywords

atherosclerosis vascular endothelial cells FGF-18 TRIM28 monocytes

Introduction

As the underlying pathology of cardiovascular and cerebrovascular diseases such as coronary heart disease and stroke, atherosclerosis remains a global health burden with persistently high incidence and mortality (Herrington et al., 2016; Johnson, 2017). Its development is a multifactorial and multistage process, originating from endothelial dysfunction and vascular inflammatory activation, which form the core pathophysiological mechanism (Ni et al., 2025; Zhuang et al., 2019). In particular, monocyte adhesion to the endothelium acts as a pivotal early trigger in this cascade (Liang et al., 2025).

Under physiological conditions, vascular endothelial cells function as a dynamic barrier, maintaining intravascular homeostasis through the regulation of vascular tone and inhibition of platelet aggregation (Theofilis et al., 2021). However, exposure to risk factors including hypercholesterolemia and hypertension leads to the oxidative modification of intimal lipoproteins, primarily low-density lipoprotein (LDL), by reactive oxygen species, consequently generating ox-LDL (Kattoor et al., 2019). ox-LDL, as a key pathogenic factor of atherosclerosis, can disrupt endothelial cell homeostasis through multiple pathways (Luquain-Costaz and Delton, 2024).ox-LDL can not only be recognized and endocytosed by clearance receptors (such as LOX-1) on endothelial cells, promoting lipid deposition and foam cell formation, but also activate the inflammatory signaling pathways of endothelial cells (Pirillo et al., 2013). This activation triggers the robust expression of adhesion molecules, notably VCAM-1 and ICAM-1. These molecules serve as ligands that bind to integrins on monocytes, mediating their firm adhesion to the endothelium (Cybulsky et al., 2001). The adherent monocytes then migrate into the subendothelial layer, where they differentiate into macrophages. These cells then engulf lipids, transforming into foam cells that form the lipid core of atherosclerotic plaques (Libby et al., 2010). Meanwhile, activated endothelial cells also secrete various pro-inflammatory factors, such as TNF-α and MCP-1, and upregulate the expression of COX-2 and promote the production of inflammatory mediators, such as PGE₂, thereby amplifying the local inflammatory response and accelerating the process of atherosclerosis (Ricciotti and FitzGerald, 2011; Weber and Noels, 2011). Therefore, developing novel strategies to effectively inhibit endothelial inflammation and monocyte adhesion is crucial for the prevention and treatment of atherosclerosis.

The fibroblast growth factor (FGF) family encompasses a class of crucial polypeptide signaling molecules (Liu et al., 2021). Beyond its well-established roles in bone development and lung morphogenesis, FGF-18 has garnered interest for its potential vascular functions. Emerging evidence indicates that FGF-18 may confer vascular protection within the cardiovascular system (Chen et al., 2023). FGF-18, known to signal through the fibroblast growth factor receptor 1 (FGFR1), has emerging roles in maintaining vascular homeostasis, but its precise function in endothelial inflammation remains unclear.

Tripartite motif-containing 28 (TRIM28) is a transcriptional regulator with versatile functions (Friedman et al., 1996). It interacts with histone modification enzymes, chromatin remodeling complexes, etc., through its multiple domains, participating in processes such as gene transcription inhibition, autophagy regulation, and DNA damage repair (Czerwińska et al., 2017; Le Douarin et al., 1996). Beyond these functions, TRIM28 has been identified as a pivotal regulator in vascular endothelium, modulating inflammatory responses and angiogenesis through its control over TNFR1, TNFR2, and VEGFR2 expression (Wang et al., 2017). To date, it is unknown if TRIM28 mediates ox-LDL-induced endothelial injury or if it constitutes a target through which FGF-18 exerts its endothelial protection.

Grounded in the aforementioned research, this study utilized human aortic vascular endothelial cells (HAVECs) subjected to ox-LDL to model endothelial inflammation. We firstly systematically assessed the effects of rhFGF-18 on the inflammatory response and monocyte adhesion. Then, the specific role and molecular mechanism of TRIM28 in this process were deeply investigated. These efforts are intended to offer novel insights and identify potential therapeutic targets for atherosclerosis prevention and treatment.

Materials and Methods

Cell culture, transduction, and treatment

HAVECs were purchased from the American Type Culture Collection and were maintained in complete Dulbecco's Modified Eagle Medium (DMEM) containing 10% fetal bovine serum (FBS), 100 U/mL penicillin, and 100 µg/mL streptomycin at 37°C and 5% CO2. THP-1 cells were propagated in RPMI-1640 with 10% FBS. ox-LDL was purchased as a ready-to-use reagent from Beijing Solarbio Science & Technology Co., Ltd. (China) and was prepared at the specified concentrations (0, 25, 50, 100 mg/L) in serum-free medium prior to application. Recombinant human FGF-18 (rhFGF-18) was purchased from PeproTech Company in the United States. To construct the TRIM28 overexpression model, when the HAVECs fusion degree reached 60%−70%, the Ad-viral TRIM28 or empty vector was transfected with Lipofectamine 3000. The efficiency of transfection was assessed by Western blot at 48 h post-transfection.

Real-Time quantitative PCR

Total RNA was extracted from cells using TRIzol reagent, and its concentration and purity were verified. Subsequently, 1 µg of RNA was reverse-transcribed into cDNA. Quantitative real-time PCR was performed using Synergy Brands (SYBR) Green chemistry with Glyceraldehyde-3-Phosphate Dehydrogenase (GAPDH) as the endogenous control. The amplification protocol included initial denaturation, 40 cycles of annealing/extension, and a final melt curve analysis. All reactions were run in triplicate, and relative gene expression was calculated using the 2^–ΔΔCt method.

Nuclear fraction isolation and Western blot analysis

After incubating HAVECs with Radioimmunoprecipitation Assay Buffer (RIPA) lysis buffer on ice for 30 min, the lysates were centrifuged at 12,800 g and 4°C for 15 min. The resulting supernatant was then collected, and total protein concentration was determined by the Bicinchoninic Acid Assay (BCA) assay. For subcellular fractionation, nuclear and cytoplasmic proteins were separated using a dedicated extraction kit. Following concentration determination by BCA, protein samples were separated by SDS-PAGE (80 V in stacking gel, 120 V in separating gel) and transferred to a PVDF membrane (100 V, 90 min). The membrane was blocked with 5% nonfat milk for 2 h at room temperature, followed by incubation with primary antibody overnight at 4°C. Subsequently, the membrane was incubated with an Horseradish Peroxidase (HRP)-conjugated secondary antibody for 1 h. Protein bands were visualized using an Enhanced Chemiluminescence (ECL) chemiluminescence kit and quantified by analyzing band intensity with ImageJ software. The following primary antibodies were used: anti-LOX-1 (Abcam, UK, ab126538), anti-COX-2 (Abcam, UK, ab179800), anti-VCAM-1 (Abcam, UK, ab134047), anti-ICAM-1 (Abcam, UK, ab171123), anti-phospho-TRIM28 (Ser824) (Thermo Fisher Scientific, 702084), and anti-TRIM28 (Thermo Fisher Scientific, MA1-2023).

Enzyme-Linked Immunosorbent Assay

The concentrations of TNF-α and MCP-1 were measured with commercially available Enzyme-Linked Immunosorbent Assay (ELISA) kits (TNF-α: Abcam, UK, ab181421; MCP-1: Abcam, UK, ab179886), and PGE₂ levels were quantified using an ELISA kit (Thermo Fisher Scientific, EHPGE2). Following collection, HAVECs from each experimental group were processed in accordance with the ELISA kit protocol. The samples were centrifuged (300g, 5 min, 4°C) to collect the supernatant. Subsequent to a 1-h primary antibody incubation and 3 Phosphate-Buffered Saline (PBS) washes, an HRP-conjugated secondary antibody was introduced for 20 min. Readings were acquired at 450 nm on a microplate reader.

Measurement of THP-1 cell adhesion to HAVECs using calcein acetoxymethyl ester (calcein AM)

Following the initial incubation, cells were thoroughly washed with PBS to remove any unbound components and carefully resuspended in fresh medium to ensure a single-cell suspension for the subsequent adhesion assay. For the coculture, pre-labeled THP-1 monocytes, at a concentration of 5 × 10⁵ cells, were introduced onto confluent monolayers of HAVECs seeded at 1 × 10⁵ cells. This coculture system was maintained for 2 h under standard culture conditions to allow for firm adhesion. After this incubation, non-adherent monocytes were meticulously removed by performing 3 sequential, gentle washes with pre-warmed PBS. The remaining adherent fluorescent cells in multiple, randomly selected fields per well were then visualized and imaged using an inverted fluorescence microscope. Finally, the number of adhered cells in each field was precisely quantified through image analysis using ImageJ software.

Statistical analysis

We performed statistical analyses using GraphPad Prism 8.0, with all quantitative data presented as mean ± standard deviation. Student’s t-test was used for two-group comparisons, while Analysis of Variance (ANOVA) with Tukey’s post hoc test was applied for multigroup comparisons, with the threshold for statistical significance set at P < 0.05.

Results

ox-LDL reduced the levels of p-FGFR1 in HAVECs

HAVECs were exposed to a range of ox-LDL concentrations (0, 25, 50, 100 mg/L) for 24 h. Western blot analysis (Fig. 1A) revealed a pronounced, dose-dependent decline in p-FGFR1 levels with increasing ox-LDL concentrations compared with the control. To assess temporal effects, cells were exposed to 100 mg/L ox-LDL for different durations (0, 12, 24, 48 h). As shown in Figure 1B, p-FGFR1 levels began to decrease significantly after 12 h, reaching a minimum at 24 h. Although a slight recovery was observed at 48 h, the level remained markedly lower than the baseline (0 h), demonstrating that the inhibition of p-FGFR1 by ox-LDL is also time-dependent.

FIG. 1.

ox-LDL reduces the levels of p-FGFR1 in human aortic vascular endothelial cells (HAVECs). (A) HAVECs were stimulated with ox-LDL (0, 25, 50, 100 mg/L) for 24 h, and the levels of p-FGFR1 were measured by western blot analysis. (B) HAVECs were stimulated with ox-LDL (100 mg/L) for 12, 24, and 48 h, and the levels of p-FGFR1 were measured by western blot analysis (N = 6; ^##, ^###, P < 0.01, 0.005 vs. control group). ox-LDL, oxidized low-density lipoprotein.

rhFGF-18 inhibits the expression of LOX-1 in HAVECs induced by ox-LDL

The modulatory effect of rhFGF-18 on ox-LDL-induced LOX-1 was examined in HAVECs subjected to combined treatment for 24 h. qPCR data showed that rhFGF-18 concentration-dependently counteracted the ox-LDL-mediated upregulation of LOX-1 mRNA (Fig. 2A). Corresponding Western blot results further demonstrated a dose-dependent reduction in LOX-1 protein levels (Fig. 2B), revealing a concerted inhibition of ox-LDL-induced LOX-1 expression by rhFGF-18 at both transcriptional and translational levels.

FIG. 2.

rhFGF-18 reduces the expression of LOX-1 induced by ox-LDL in human aortic vascular endothelial cells (HAVECs). HAVECs were stimulated with ox-LDL (100 mg/L) in the presence or absence of rhFGF-18 (25, 50 ng/mL) for 24 h. (A) LOX-1 mRNA levels; (B) LOX-1 protein levels (^^^, P < 0.005 vs. control group; ^##, ^###, P < 0.01, 0.005 vs. ox-LDL group).

rhFGF-18 inhibited the expression of pro-inflammatory mediators in HAVECs

We investigated whether rhFGF-18 modulates the ox-LDL-induced expression of the key pro-inflammatory mediators TNF-α and MCP-1 by quantifying their mRNA and protein levels. The qPCR results (Fig. 3A) showed that, compared with the control group, the mRNA levels of TNF-α and MCP-1 in the ox-LDL alone treatment group were significantly increased; after combined treatment with rhFGF-18, the mRNA levels of both were significantly decreased. The ELISA results further verified the changes in protein levels (Fig. 3B): Relative to the control, ox-LDL stimulation notably increased TNF-α and MCP-1 concentrations to 26.2 ± 2.99 and 46.9 ± 5.82 pg/mL, respectively. Co-treatment with 20 and 50 ng/mL rhFGF-18 notably attenuated this increase. The higher concentration of 50 ng/mL effectively suppressed the levels of pro-inflammatory mediators, reducing TNF-α and MCP-1 to 14.6 ± 1.52 and 22.8 ± 3.09 pg/mL, respectively.

FIG. 3.

rhFGF-18 inhibits the expression of pro-inflammatory mediators in human aortic vascular endothelial cells (HAVECs). HAVECs were stimulated with ox-LDL (100 mg/L) with or without rhFGF-18 (25, 50 ng/mL) for 24 h. (A) mRNA levels of TNF-α and MCP-1 measured by real-time PCR; (B) Protein levels of TNF-α and MCP-1 measured by ELISA (^^^, P < 0.005 vs. control group; ^##, ^###, P < 0.01, 0.005 vs. ox-LDL group). ELISA, Enzyme-Linked Immunosorbent Assay.

rhFGF-18 suppressed the expression of COX-2 and the production of PGE₂ in HAVECs

Analysis of the COX-2/PGE₂ axis demonstrated that ox-LDL stimulation induced a concomitant increase in both COX-2 mRNA (3.3 ± 0.37-fold) and protein (3.0 ± 0.33-fold) expression levels relative to the control group. This induction was markedly suppressed by 25 and 50 ng/mL rhFGF-18, with 50 ng/mL rhFGF-18 reducing the levels of COX-2 mRNA and protein to 1.7 ± 0.18-fold and 1.5 ± 0.14-fold, respectively (Fig. 4A and B). Concomitantly, ELISA results showed that the ox-LDL-induced increase in PGE₂ content (139.5 ± 14.65 pg/mL) was significantly attenuated by 25 and 50 ng/mL rhFGF-18 treatment, decreasing to 101.5 ± 11.8 and 73.6 ± 8.23 pg/mL (Fig. 4C), respectively. Taken together, our findings reveal a potent inhibitory effect of rhFGF-18 on ox-LDL-triggered endothelial inflammation, significantly reducing both pro-inflammatory mediator expression and PGE₂ production.

FIG. 4.

rhFGF-18 suppresses the expression of COX-2 and the production of PGE₂ in HAVECs. HAVECs were stimulated with ox-LDL (100 mg/L) and rhFGF-18 (25, 50 ng/mL) for 24 h. (A) COX-2 mRNA levels; (B) COX-2 protein levels; (C) PGE₂ production measured by ELISA (^^^, P < 0.005 vs. control group; ^##, ^###, P < 0.01, 0.005 vs. ox-LDL group).

rhFGF-18 suppressed the expression of cell adhesion molecules VCAM-1 and ICAM-1

As revealed by qPCR, ox-LDL exposure induced a pronounced upregulation of adhesion molecule mRNA, with VCAM-1 and ICAM-1 levels being notably elevated to 3.4 ± 0.33-fold and 3.9 ± 0.41-fold relative to controls, respectively. This induction was suppressed by rhFGF-18 in a dose-dependent manner, with 50 ng/mL reducing the expression to 1.7 ± 0.17-fold and 1.9 ± 0.19-fold (Fig. 5A).

FIG. 5.

rhFGF-18 suppresses the expression of cell adhesion molecules VCAM-1 and ICAM-1. HAVECs were stimulated with ox-LDL (100 mg/L) and rhFGF-18 (25, 50 ng/mL) for 24 h. (A) mRNA levels of VCAM-1 and ICAM-1; (B) Protein levels of VCAM-1 and ICAM-1 (^^^, P < 0.005 vs. control group; ^##, ^###, P < 0.01, 0.005 vs. ox-LDL group).

A parallel trend was observed at the protein level by Western blot (Fig. 5B): the ox-LDL-induced increases in VCAM-1 (256.3 ± 29.38 pg/mg protein) and ICAM-1 (358.9 ± 43.77 pg/mg protein) were significantly attenuated by rhFGF-18, decreasing to 145.6 ± 16.76 and 198.6 ± 23.55 pg/mg protein, respectively.

rhFGF-18 reduces the adhesion of THP-1 monocytes to HAVECs induced by ox-LDL

To evaluate monocyte-endothelial adhesion, we employed the Calcein AM staining assay. Fluorescence imaging revealed a substantial increase in adhered THP-1 cells following ox-LDL treatment compared with the control, which was markedly attenuated by co-treatment with 25 and 50 ng/mL rhFGF-18 in a dose-dependent manner (Fig. 6A). Quantitative analysis showed that the relative adhesion rate was 2.9 ± 0.28 times in the ox-LDL alone treatment group and decreased to 2.1 ± 0.22 and 1.5 ± 0.16 times in the 25 and 50 ng/mL rhFGF-18 group (Fig. 6B).

FIG. 6.

rhFGF-18 reduces the adhesion of THP-1 monocytes to ox-LDL-induced HAVECs. HAVECs were challenged with ox-LDL (100 mg/L) and rhFGF-18 (25, 50 ng/mL) for 24 h. THP-1 monocyte adhesion was assessed using calcein AM staining. (A) Representative images of THP-1 monocyte adhesion. Scale bar, 50 μm; (B) Quantification of adherent THP-1 monocytes (^^^, P < 0.005 vs. control group; ^##, ^###, P < 0.01, 0.005 vs. ox-LDL group). ox-LDL, oxidized low-density lipoprotein.

rhFGF-18 prevented the phosphorylation and nuclear translocation of TRIM28

The phosphorylation and nuclear translocation of TRIM28 were examined by Western blot. Results demonstrated that ox-LDL treatment significantly increased the level of p-TRIM28 to 3.2 ± 0.34-fold of the control (Fig. 7A), and the level of TRIM28 in the nucleus was 2.4 ± 0.25 times (Fig. 7B). After treatment with 25 and 50 ng/mL rhFGF-18, p-TRIM28 decreased to 2.1 ± 0.23 and 1.2 ± 0.14 times, and nuclear TRIM28 decreased to 1.7 ± 0.18 and 1.3 ± 0.11 times.

FIG. 7.

rhFGF-18 inhibits the phosphorylation and nuclear translocation of TRIM28. HAVECs were challenged with ox-LDL (100 mg/L) and rhFGF-18 (25, 50 ng/mL) for 24 h. (A) Levels of p-TRIM28; (B) Nuclear levels of TRIM28 (^^^, P < 0.005 vs. Control group; ^##, ^###, P < 0.01, 0.005 vs. ox-LDL group).

Overexpression of TRIM28 abolishes the anti-inflammatory and anti-adhesive effects of rhFGF-18 in HAVECs

To elucidate the necessity of TRIM28 in rhFGF-18-mediated protection, we established TRIM28-overexpressing HAVECs (Fig. 8A). Results demonstrated that in empty vector control cells, rhFGF-18 effectively suppressed the ox-LDL-induced upregulation of VCAM-1 and ICAM-1. Conversely, in TRIM28-overexpressing cells, the protective effect of rhFGF-18 was completely abolished (Fig. 8B). Furthermore, the Calcein-AM adhesion assay revealed that TRIM28 overexpression abolished the inhibitory effect of rhFGF-18 on THP-1 monocyte adhesion to HAVECs induced by ox-LDL (Fig. 8C), functionally confirming that TRIM28 is essential for the anti-adhesive action of rhFGF-18.

FIG. 8.

Overexpression of TRIM28 abolishes the anti-inflammatory and anti-adhesive effects of rhFGF-18 in HAVECs. HAVECs were transduced with Ad-viral TRIM28 (TRIM28 OE) or an empty vector, followed by stimulation with ox-LDL (100 mg/L) and/or rhFGF-18 (50 ng/mL) for 24 h. (A) Western blot analysis confirming TRIM28 overexpression; (B) Protein levels of VCAM-1 and ICAM-1. (C) Quantification of adherent THP-1 monocytes (^^^, P < 0.005 vs. control group; ^##, ^###, P < 0.01, 0.005 vs. ox-LDL group).

Discussion

Atherosclerosis prevention and treatment continue to be a major challenge in cardiovascular medicine. Endothelial inflammation and monocyte adhesion represent critical early steps in disease pathogenesis. Given their established role, elucidating the underlying regulatory mechanisms holds considerable therapeutic potential for the development of novel therapeutic interventions. The research systematically explored the role and mechanism of rhFGF-18 in ox-LDL-induced endothelial inflammation. It was confirmed for the first time that rhFGF-18 can alleviate endothelial inflammation and reduce monocyte adhesion by inhibiting TRIM28 activation, offering a novel theoretical framework for the work of the mechanism of atherosclerosis.

This study first discovered that ox-LDL down-regulated the level of p-FGFR1 in HAVECs in a dose- and time-dependent manner. FGFR1 is the main functional receptor of FGF-18, and its phosphorylation level directly reflects the activation status of the FGF-18/FGFR1 signaling pathway. As a core pathogenic factor of atherosclerosis, the inhibitory effect of ox-LDL on p-FGFR1 suggests that ox-LDL may disrupt vascular endothelial homeostasis by interfering with the normal activation of the FGF-18/FGFR1 signaling pathway, thereby inducing inflammatory responses. This evidence corroborates the inhibitory effect of ox-LDL on protective signaling pathways, such as PI3K/Akt, in previous studies (Huo et al., 2022). By activating the FGFR1 signaling pathway, exogenous rhFGF-18 can thereby antagonize the detrimental effects of ox-LDL. This finding lays the groundwork for subsequent validation of the protective role of rhFGF-18.

LOX-1, a key ox-LDL clearance receptor on vascular endothelial cells, mediates the internalization of ox-LDL, leading to the subsequent activation of downstream inflammatory signaling. Studies have shown that, in mice with LOX-1 gene knockout, the thickening of the arterial intima is alleviated, inflammation is weakened, and the expression of protective factors is upregulated. However, overexpression of LOX-1 significantly accelerates the formation of atherosclerotic lesions and is accompanied by more severe inflammatory responses (Pirillo et al., 2013). Our data demonstrate that rhFGF-18 concentration-dependently suppresses the ox-LDL-induced upregulation of LOX-1 in HAVECs. This suggests that rhFGF-18 may exert its protective effects via an upstream mechanism: by downregulating LOX-1 expression, it potentially reduces the cellular uptake and subsequent accumulation of ox-LDL, thereby attenuating the inflammatory injury initiated by this ligand.

The excessive production of pro-inflammatory mediators is an important feature of endothelial inflammatory responses and a key factor promoting the development of atherosclerosis. TNF-α, as a classical proinflammatory factor, can amplify inflammatory responses by activating pathways such as NF-κB (Aggarwal, 2003). MCP-1 is a key factor for monocyte chemotaxis, which can specifically attract monocytes to migrate to the inflammatory site and adhere to endothelial cells (Pola et al., 2004). The upregulation of COX-2, a pivotal enzyme in prostaglandin synthesis, leads to excessive formation of mediators such as PGE₂, further aggravating inflammation (Martín-Vázquez et al., 2023). This study revealed that rhFGF-18 broadly suppresses ox-LDL-induced expression of TNF-α, MCP-1, and COX-2 (mRNA and protein) in HAVECs, and correspondingly lowers PGE₂ levels. These findings collectively indicate that rhFGF-18 mitigates endothelial inflammation by multi-targetedly inhibiting the generation and release of pro-inflammatory mediators. The observed anti-inflammatory effect is consistent with previous reports in other disease models, such as the attenuation of Lipopolysaccharide (LPS)-induced pulmonary inflammation by FGF-18 via inhibition of the NF-κB pathway (Hu et al., 2024).

VCAM-1 and ICAM-1 are critically involved in monocyte-endothelial adhesion, a fundamental event in the inflammatory initiation of atherosclerosis. VCAM-1 mainly binds to VLA-4 on the surface of monocytes, while ICAM-1 binds to LFA-1 on the surface of monocytes. Both jointly mediate the adhesion between monocytes and endothelial cells (Bei et al., 2023). Our data demonstrate that rhFGF-18 significantly suppresses the ox-LDL-induced expression of VCAM-1 and ICAM-1 in HAVECs. This molecular-level suppression was functionally confirmed by Calcein AM staining, which showed that rhFGF-18 concomitantly reduces the adhesion of THP-1 monocytes to HAVECs. These findings indicate that rhFGF-18 can interfere with the monocyte-endothelial adhesion process by downregulating key adhesion molecules, thereby directly impeding an early critical step in atherogenesis. This mechanism aligns with reports on other vasculoprotective factors, such as Vascular Endothelial Growth Factor (VEGF), which also attenuates monocyte adhesion by suppressing adhesion molecule expression (Min et al., 2005).

We focused on the transcriptional regulator TRIM28 to explore the mechanism of rhFGF-18-mediated protection. TRIM28 regulates gene expression via interaction with Krüppel-Associated Box (KRAB) zinc-finger proteins or direct modulation of transcription factors (Grassi et al., 2019). This study revealed that rhFGF-18 inhibits the ox-LDL-induced phosphorylation and nuclear translocation of TRIM28 in HAVECs. Furthermore, TRIM28 overexpression abrogated the suppressive effects of rhFGF-18 on VCAM-1 and ICAM-1 induction, demonstrating a pivotal role for TRIM28 inhibition in the protective action of rhFGF-18.

This study also has certain limitations: Primarily, the experimental work was conducted solely in an in vitro HAVEC model and lacks validation through in vivo animal studies. In the future, a mouse model of ApoE^–/– atherosclerosis needs to be constructed to further verify the in vivo effect of the FGF-18/TRIM28 axis. Second, the inflammation-related genes directly regulated downstream of TRIM28 have not been deeply explored. It is necessary to clarify their target genes and improve the regulatory network through technologies such as ChIP-seq. In addition, the usage concentration of rhFGF-18 in the study was relatively high, and the dosage and administration method need to be further optimized to offer robust evidence for evidence-based clinical decision-making.

In summary, our work demonstrates that FGF-18 alleviates ox-LDL-triggered endothelial inflammation and monocyte-endothelial adhesion, identifying the inhibition of TRIM28 activation as a key mechanism. This newly uncovered FGF-18/TRIM28 pathway offers a novel potential therapeutic strategy for combating atherosclerosis.

Data Availability Statement

The data that support the findings of this study are available from the corresponding author upon reasonable request.

Authors’ Contributions

C.Y.: Conceptualization, methodology, validation, supervision, project administration, and writing—review and editing. Z.L.: Conceptualization, methodology, project administration, and writing—review and editing. W.Y.: Investigation, formal analysis, data curation, and writing—original draft. H.D.: Investigation, resources, visualization, and writing—original draft.

Footnotes

Author Disclosure Statement

The authors declare that they have no conflicts of interest.

Funding Information

This work was supported by the hospital-level scientific research and technological breakthrough project of the First People’s Hospital of Baiyin City (No.2022YK-13).

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, et al. Endothelial Foxp1 Suppresses Atherosclerosis via Modulation of Nlrp3 Inflammasome Activation. Circ Res, 2019; 125(6):590–605.

Fibroblast Growth Factor-18 (FGF-18) Attenuates ox-LDL-Induced Endothelial Inflammation and Monocyte Adhesion by Inhibiting TRIM28

Abstract

Keywords

Introduction

Materials and Methods

Cell culture, transduction, and treatment

Real-Time quantitative PCR

Nuclear fraction isolation and Western blot analysis

Enzyme-Linked Immunosorbent Assay

Measurement of THP-1 cell adhesion to HAVECs using calcein acetoxymethyl ester (calcein AM)

Statistical analysis

Results

ox-LDL reduced the levels of p-FGFR1 in HAVECs

rhFGF-18 inhibits the expression of LOX-1 in HAVECs induced by ox-LDL

rhFGF-18 inhibited the expression of pro-inflammatory mediators in HAVECs

rhFGF-18 suppressed the expression of COX-2 and the production of PGE2 in HAVECs

rhFGF-18 suppressed the expression of cell adhesion molecules VCAM-1 and ICAM-1

rhFGF-18 reduces the adhesion of THP-1 monocytes to HAVECs induced by ox-LDL

rhFGF-18 prevented the phosphorylation and nuclear translocation of TRIM28

Overexpression of TRIM28 abolishes the anti-inflammatory and anti-adhesive effects of rhFGF-18 in HAVECs

Discussion

Data Availability Statement

Authors’ Contributions

Footnotes

Author Disclosure Statement

Funding Information

References

rhFGF-18 suppressed the expression of COX-2 and the production of PGE₂ in HAVECs