1 Introduction
Psoriasis is a chronic, recurrent inflammatory skin disease characterized by epidermal hyperkeratosis and hypokeratosis, keratinocyte proliferation, dermal microvessel dilation, and infiltration of inflammatory cells [
1,
2]. Angiogenesis, driven by cytokines like IL-17, IL-8, tumor necrosis factor α (TNF-α), angiopoietin, and vascular endothelial growth factor (VEGF), plays a key role in psoriasis development [
3]. VEGF, produced by skin keratinocytes, is the primary mediator of angiogenesis under both normal and pathological conditions by binding to vascular endothelial growth factor receptors (VEGFRs) on endothelial cells [
3]. In psoriasis, VEGF levels are notably higher in psoriatic lesions compared to normal skin and are linked to disease severity [
4,
5]. Inflammatory conditions increase microvascular permeability and stimulate endothelial cell division and proliferation, enhancing angiogenesis [
6]. Moreover, the Th17 cells and the IL-23/IL-17 pathway are crucial in psoriasis pathogenesis, with IL-17 from Th17 cells directly stimulating angiogenesis [
7,
8]. Elevated IL-17 levels in psoriasis can also trigger vascular inflammation [
9] and boost pro-inflammatory factors [
10–
12], leading to further angiogenesis [
13,
14].
N
6-methyladenosine (m
6A), a dynamic mRNA modification at the sixth N position of adenine, typically occurs every 85 nucleotides, particularly enriching the coding sequence (CDS), 3′ untranslated region (3′UTR), and stop codon region [
15–
18]. Found in eukaryotic and viral mRNAs, m
6A regulates post-transcriptional processes [
19], and m
6A methylation impacts RNA splicing, transport, and stability [
20]. The modification process is regulated by three proteins, including methyltransferases (writers), demethyltransferases (erasers), and binding proteins (readers). In mammals, methyltransferase-like 3 (METTL3), METTL14, and Wilms tumor 1-associated protein (WTAP) are the primary m
6A methyltransferases, while fat mass and obesity-associated protein (FTO) and AlkB homolog 5 (ALKBH5) are the demethylases. m
6A binding proteins, including YTH N
6-methyladenosine RNA binding protein 1 (YTHDF1), YTHDF2, and YTH domain-containing protein 1 (YTHDC1), manage RNA functions in the nucleus [
21].
ALKBH5, an RNA demethylase, modulates cellular processes by altering m
6A demethylation, impacting RNA regulation and gene expression crucial for cancer progression and inflammation-related disorders [
22–
27]. Studies have shown that ALKBH5 knockdown reduces proliferation, migration, and angiogenesis in lung cancer cells [
28], and its silencing decreases UVB-induced proliferation and inflammation in HaCaT cells [
29]. Inhibition of ALKBH5 also limits skin cell proliferation and inflammation at wound edges [
30]. Conversely, its overexpression in retinal pigment epithelium cells boosts VEGF-A levels and AKT phosphorylation [
31], while AKT/mTOR pathway activation is linked to autophagy, inflammation, and angiogenesis in psoriasis [
32]. Furthermore, altering ALKBH5 expression affects cardiac endothelial cell functions including proliferation, migration, and tube formation [
33]. Despite its established role in m
6A-related gene regulation in psoriasis vulgaris, research on ALKBH5’s impact on psoriasis inflammation and angiogenesis remains limited [
34–
36].
Further investigation of ALKBH5’s role in m6A modification is crucial for understanding psoriasis pathogenesis, a condition linked to inflammation and angiogenesis. This study shows high ALKBH5 expression in psoriatic lesions in IMQ mice and an IL-17A-induced human umbilical vein endothelial cells (HUVECs) angiogenesis model, suggesting that ALKBH5-mediated demethylation may influence psoriasis development, providing new insights into its mechanisms.
2 Materials and methods
2.1 Reagent preparation
IOX1, sourced from Selleck (S7234, USA), was dissolved in a solution of 10% DMSO (from Sigma), 20% ethanol, Cremophor EL (1:1 ratio), and 70% sterile saline. The experimental group of mice received daily injections of IOX1 at doses of 5, 10, or 20 mg/kg, with a volume of 90 μL per mouse. The model and control groups received the same volume of solvent daily.
2.2 Animal source and feeding
The study utilized 6–8-week-old male C57BL/6 wild-type mice purchased from the Medical Laboratory Animal Center (Guangdong, China). Alkbh5+/− mice (S-KO-08566) on C57BL/6Ncya background were obtained from Cyagen Biosciences Inc. Alkbh5−/− mice and C57BL/6N wild-type mice of the littermates were generated through in vitro fertilization at Shulaibao Biotechnology Co., Ltd. (Wuhan, China). Institutional Animal Care and Use Committee of Shenzhen Top Biotech Co., Ltd approved all animal procedures, which followed ARRIVE guidelines.
2.3 Imiquimod induced inflammation in a psoriasis-like mouse model
After depilating the posterior neck skin, mice were grouped (n = 5 per group). The experimental group received 62.5mg of 5% imiquimod cream (Mingxin, Sichuan, China) and a hypodermic injection of IOX1. The model group was treated similarly with imiquimod cream and an isodose solvent injection, while the control group received an equivalent dose of vaseline and solvent injection. Treatments lasted for 5 days, with euthanasia and assessments on the 6th day. Mice were weighed and photographed daily. The severity of inflammation was evaluated using the Psoriasis Area and Severity Index (PASI), scoring erythema, scaling, and thickening on a 0–4 scale (0 = none; 1 = slight; 2 = moderate; 3 = marked; 4 = highly marked), with a total possible score of 0–12. PASI score was evaluated by an evaluator independent of the inhibitor dose.
2.4 Isolating epidermal and dermal layers from mice skin
Posterior neck skin from both the IMQ and control groups was excised, rinsed thrice with PBS, cut into 2 cm × 2 cm pieces, and incubated in 0.25% dispase at 37 °C for one hour. The epidermis and dermis were separated with tweezers, and the isolated tissues were stored at −80 °C for future experiments.
2.5 Histology and immunohistochemistry
The mice’s neck skin tissue was fixed in 4% paraformaldehyde, embedded in paraffin, and sliced into 15 μm sections, which were partially stained with hematoxylin and eosin. Epidermal thickness was measured across four random fields per mouse using ImageJ software. For immunohistochemistry, paraffin sections were deparaffinized, hydrated with alcohol, and treated with Tris-EDTA antigen retrieval solution (pH 9.0, Servicebio). Sections were blocked with 3% bovine serum albumin for 30 min at room temperature, then incubated overnight at 4 °C with primary antibodies ALKBH5 (1:1000, Abcam), VEGF (1:200, Servicebio), and VEGFR (1:200, Servicebio). They were subsequently exposed to HRP-conjugated goat anti-rabbit IgG (1:200, Servicebio) for 1 h at room temperature. DAB kit (Servicebio) was used for color development, followed by nuclei counterstaining with hematoxylin (Servicebio). After dehydration and sealing with neutral gum, positive staining was quantified in three fields per section under a microscope by two independent experimenters.
2.6 Immunofluorescence
Mouse skin paraffin sections underwent deparaffinization, alcohol hydration, antigen retrieval, and PBS washing. They were then treated with 0.3% TritonX-100 for 1 h at room temperature, followed by blocking with 5% BSA for another hour at room temperature. Co-staining was performed by incubating tissue overnight at 4 °C with primary antibodies ALKBH5 (1:1000, Abcam) and CD31 (1:1000, Abcam). Fluorescent secondary antibodies corresponding to the primary antibodies were then incubated in the dark at room temperature for 1 h. The fluoresceins used were FITC (1:1000, Abcam) and Alexa Fluor® 647 (1:1000, Abcam). Nuclear staining was done with antifade mounting solution containing DAPI for 30 min before imaging. Images were captured at 3–4 random spots per skin section at 20× magnification.
2.7 HUVEC cell culture and treatment
HUVECs were sourced from Pricella Biotechnology Co., Ltd. (Wuhan, China) and cultured at 37 °C in a 5% CO2 incubator with 70%–80% humidity. The medium included 5% fetal bovine serum (Gibco, USA), 1% endothelial cell growth factor (ECGF), and 1% penicillin-streptomycin. HUVECs (1.5 × 105 cells/well) were seeded into 6-well plates and exposed to 20 ng/mL recombinant human IL-17A (MedChemExpress, USA) for 48 h to establish an in vitro angiogenesis model. Gemma Genes (Shanghai, China) synthesized small interfering RNA (siRNA) targeting ALKBH5 and an ALKBH5 overexpression plasmid, which were transfected using jetPRIME reagent (Polyplus, France). For transfection, 5 μL of siRNA (plasmid) was diluted in 200 μL of jetPRIME buffer, followed by the addition of 4 μL of jetPRIME reagent and incubation at room temperature for 10 min. Transfection efficiency was evaluated by Western blotting 48 h post-transfection. The sequences for siALKBH5, si-NC, and GAPDH primers used in knockdown experiments are listed in Table S1.
2.8 Western blotting
Mouse skin tissue and HUVEC protein samples were lysed in RIPA buffer with 1% protease/phosphatase inhibitors. Protein concentration was measured using the BCA kit. Proteins were separated by 10% SDS-PAGE and transferred to PVDF membranes. Membranes were blocked with 5% skim milk powder for 1 h at room temperature, then incubated overnight at 4 °C with primary antibodies, including VEGFA, VEGFR2 (1:1000, Abclonal); AKT, phosphor-AKT (p-AKT), mTOR, phospho-mTOR (p-mTOR) (1:1000, Cell Signaling Technology); ALKBH5 (1:1000, Abcam) and β-actin (1:1000, Servicebio). They were then treated with a horseradish peroxidase-labeled goat anti-rabbit IgG secondary antibody (1:10000) for 1 h at room temperature. Protein bands were visualized using the ECL chemiluminescence kit, and expression levels were analyzed using ImageJ software, with β-actin serving as the internal control.
2.9 Reverse transcription-quantitative PCR
RNA was extracted from mouse skin tissues or HUVECs using Trizol (Vazyme) and assessed with NanoDrop (Thermo Fisher Scientific). The RNA was then reversely transcribed into cDNA using a kit (ABclonal). Quantitative PCR was performed with the SYBR Green PCR kit (Servicebio). The Applied Biosystems StepOne™ System analyzed the results, and relative gene expression levels were calculated using the 2–ΔΔCt method. Sangon Biotech Co (Shanghai, China) designed and synthesized the qPCR primers, with sequences listed in Table S2. β-actin was used as the internal control.
2.10 Cell proliferation
HUVECs were seeded in 96-well plates at 3 × 103 cells/well with 100 µL of culture medium and incubated at 37 °C for 24 h until 80% confluence. After 48 h of siRNA or plasmid transfection, with or without IL-17A, 10 µL of CCK-8 solution was added and incubated at 37 °C for 3 h. The absorbance (OD) at 450 nm was then measured to determine cell proliferation. Cell proliferation rate = (absorption value of experimental group – absorption value of blank control)/(absorption value of control group – absorption value of blank control) × 100%.
2.11 Tube formation assay
Matrigel (Corning) was thawed overnight at 4 °C. Pre-cooled pipette tips and 24-well plates were used. Matrigel (220 µL/well) was incubated at 37 °C for 1 h. HUVECs (5 × 105 cells/mL) were seeded into the Matrigel-coated wells and incubated at 37 °C with 5% CO2 for 6 h. Capillary-like structures were observed and captured under an inverted microscope, and ImageJ software was used to quantify nodes, meshes, junctions, and branch lengths.
2.12 Statistical analysis
The results were shown as mean ± standard deviation (SD). All data were analyzed using GraphPad Prism 7.0. One-way ANOVA was used for comparing multiple groups, and t-test was applied for comparing two groups. P < 0.05 was considered statistically significant, denoted as *P < 0.05, **P < 0.01, ***P < 0.001, ****P < 0.0001.
3 Results
3.1 ALKBH5 expression was upregulated in skin lesions of IMQ-induced psoriasis mouse model, particularly in endothelial cells
To investigate the role of ALKBH5 in the development and progression of psoriasis, an IMQ-induced psoriasis mouse model was established. Western blotting analysis revealed significantly higher levels of ALKBH5 in the IMQ group than in the control group (Fig.1 and 1B). Immunofluorescence staining on skin samples showed a notable increase in ALKBH5 expression in the IMQ group, especially in dermal blood vessels. Histological staining displayed the typical linear or punctate morphology of these vessels (Fig. S1A). The immunofluorescence staining analysis provided evidence of the co-expression of ALKBH5 and CD31, a marker associated with angiogenesis, within the endothelial cells of the superficial dermis in mice skin. Notably, this co-expression was more pronounced in the group treated with IMQ (Fig.1). Furthermore, Western blotting analysis of mice dermis tissue showed that ALKBH5 and CD31 protein levels were significantly higher in the IMQ group compared to the CTL group, with a time-dependent increase (Fig. S1B). Overall, these findings indicated that ALKBH5 is prominently upregulated in psoriatic lesions in mice, with a predominant presence in endothelial cells within the superficial dermis.
3.2 ALKBH5 inhibitor IOX1 alleviated inflammation and angiogenesis in the IMQ-induced psoriasis mouse model
Next, IOX1 was employed to further investigate a potential role of ALKBH5 in psoriasis. C57BL/6 wild mice were divided into three groups: control, IMQ-treated, and IMQ-treated with varying concentrations (5, 10, or 20 mg/kg/day) of IOX1 groups [
37]. The experimental design and drug administration details are outlined in Fig.2. Immunohistochemistry and Western blotting analysis confirmed that IOX1 downregulated ALKBH5 expression in skin lesions compared with the IMQ group (Fig.2 and 2C). Furthermore, IOX1 significantly improved psoriatic inflammation in mice, such as skin thickening, scaling, and erythema, with a dose-dependent manner (Fig.2). H&E staining showed slight hyperkeratosis and inflammatory cell infiltration in the IOX1 group compared with the IMQ group (Fig.2). On the 6th day, the IOX1 group showed a significant reduction in PASI scores and epidermal thickness measurements compared with the IMQ group (Fig.2).
ALKBH5 co-localizes with CD31 in skin lesions of psoriasis-like mice, indicating a potential role in angiogenesis. Vascular photographs of mouse skin revealed a significant decrease in angiogenesis in the IOX1-treated group, indicating the therapeutic potential of ALKBH5 in this pathological process (Fig.2). Additionally, the expression of VEGF, a crucial factor in psoriatic angiogenesis, was significantly reduced in the lesions of IOX1-treated mice, as demonstrated by immunohistochemistry (Fig.2). Taken together, these data suggested that IOX1, acting as an inhibitor of ALKBH5, effectively suppressed abnormal angiogenesis induced by IMQ and reduced skin inflammation in psoriasis-like mice.
3.3 ALKBH5-KO alleviated psoriasis-like dermatitis and angiogenesis
To further investigate the role of ALKBH5 in vivo, we utilized ALKBH5-KO mice, revealing its impact on psoriasis pathogenesis. Immunohistochemistry and Western blotting analysis confirmed successful ALKBH5 knockout in skin lesions (Fig.3 and 3B), leading to a significant decrease in ALKBH5 levels. ALKBH5-KO mice exhibited marked improvements in IMQ-induced psoriatic inflammation and histological changes in the ALKBH5-KO group compared with WT littermates (Fig.3). Furthermore, statistically significant differences in PASI scores between WT and ALKBH5-KO mice first emerged on day 5, while epidermal thickening significantly decreased in the ALKBH5-KO mice on day 6 (Fig.3). Meanwhile, mRNA expression levels of pro-inflammatory mediators IL-17A, IL-1β, and CXCL12 in skin lesions were reduced in the ALKBH5-KO mice compared with the WT littermates (Fig.3).
Subsequently, the impact of angiogenesis in psoriasis-like lesions was further elucidated, and clinical features revealed a notable decrease in blood vessel formation in the skin lesions of ALKBH5-KO mice compared with WT littermates. Meanwhile, immunohistochemistry analysis showed reduced expression of VEGF and VEGFR in the skin lesions of ALKBH5-KO mice (Fig.3). Furthermore, the colocalization of ALKBH5 and CD31 expression was examined in the control, IMQ, and ALKBH5-KO + IMQ groups, revealing a significant decrease in colocalization of ALKBH5 and CD31 in the ALKBH5-KO mice compared with WT littermates (Fig.3). Consistent with these findings, Western blotting results demonstrated protein expression of VEGFA and VEGFR2 downregulated in the skin lesions of ALKBH5-KO mice. Given the role of the AKT/mTOR pathway in regulating angiogenesis in psoriasis, Western blotting analysis of AKT/mTOR pathway-related proteins in mouse skin lesions showed reduced protein expression of p-mTOR and p-AKT after ALKBH5-KO (Fig.3). Overall, Alkbh5 deficiency significantly improved IMQ-induced psoriasis-like mice skin inflammation and angiogenesis compared with WT mice, its potential anti-inflammatory mechanisms may rely on the modulation of the AKT-mTOR pathway.
3.4 IL-17A stimulation upregulated the expression of ALKBH5 in HUVECs
Endothelial cells play a crucial role in the increased angiogenesis observed in psoriasis. The pathogenesis of psoriasis involves the IL-23–Th17–IL-17 axis. Thus, we determined whether IL-17A could enhance the expression of ALKBH5 in HUVECs. Western blotting results demonstrated that exposure to 20 ng/mL of IL-17A for 24 h and 48 h significantly increased the expression of ALKBH5 protein levels compared with the normal control group (Fig.4). RT-qPCR analysis revealed that IL-17A stimulation upregulated the mRNA expression of ALKBH5, IL-17RA, CD31, and VEGF in HUVECs (Fig.4 to 4E). Furthermore, Western blotting analysis indicated that exposure to certain concentration (20 ng/mL) of IL-17A for 48 h significantly upregulated the expression of ALKBH5 as well as angiogenesis-related proteins VEGFA and VEGFR2 (Fig.4 and 4G). Altogether, these findings suggested that IL-17A promoted ALKBH5 expression to enhance angiogenesis. The CCK-8 assay also demonstrated that treatment with IL-17A resulted in a concentration-dependent effect which increased HUVEC proliferation at 24 h and 48 h (Fig.4).
To investigate the involvement of other cytokines in regulating the expression of ALKBH5 in HUVECs, the cells were treated with psoriatic inflammatory cytokines IL-6 (20 ng/mL) or IL-23 (50 ng/mL) for 24 h (Fig. S2A and S2B), and TNF-α (10 ng/mL) for 24 h and 48 h, respectively (Fig. S2C). However, no difference in the expression of ALKBH5 protein was observed in HUVECs. Meanwhile, stimulation with M5 (2.5 ng/mL) for 48 h, as well as treatment with IL-17A at various concentrations and time gradients in HaCat cells could not find any difference in the expression of ALKBH5 protein (Fig. S2D–S2F). These experiments reconfirmed the previous results of immunofluorescence in vivo, ALKBH5 was specifically expressed on endothelial cells.
3.5 ALKBH5 knockdown inhibited whereas ALKBH5 overexpression promoted endothelial cell proliferation
The process of angiogenesis stimulates the VEGF activation, to further promotes migration, and proliferation of endothelial cells, which playing a crucial role in psoriasis-like inflammation. VEGFR2 serves as the primary recognition receptor. Both siRNA knockdown and plasmid overexpression of ALKBH5 were utilized to investigate the regulatory impact of ALKBH5 on HUVEC cell proliferation. The efficiency of knockdown and overexpression was proved through Western blotting analysis (Fig.5 and 5B), with siALKBH5-1076 showing more pronounced knockdown effects at 48 h and thus it was selected for subsequent experiments. RT-qPCR analysis revealed that siALKBH5 decreased IL-17RA expression (Fig.5 and 5D), while ALKBH5 overexpression increased IL-17RA levels (Fig.5 and 5F). Notably, the mRNA level of ALKBH5 and IL-17RA showed no significant differences after IL-17A stimulation, possibly due to insufficient IL-17A dose. The regulatory impact of ALKBH5 expression on HUVEC cell proliferation was also assessed. ALKBH5-knockdown inhibited HUVEC cell proliferation in the presence of IL-17A (20 ng/mL) for 48 h. Conversely, ALKBH5 overexpression dramatically promoted cell proliferation (Fig.5 and 5H). Together, these findings indicate that ALKBH5 plays a role in regulating vascular endothelial cell proliferation.
3.6 ALKBH5 knockdown inhibited while ALKBH5 overexpression promoted endothelial cell tube formation
Microvascular expansion is a primary clinical characteristic of psoriasis. Our previous findings suggested that IL-17A not only promoted angiogenesis in endothelial cells but also induced ALKBH5 expression in HUVECs. Hence, we conducted Western blotting to investigate the impact of regulating ALKBH5 expression on angiogenesis in HUVECs. The results indicated that downregulation of ALKBH5 decreased the levels of angiogenesis-related proteins VEGFA and VEGFR2 (Fig.6), while upregulation of ALKBH5 increased VEGFA and VEGFR2 expression (Fig.6). To further assess whether regulating ALKBH5 expression in HUVECs has an effect on angiogenesis, we used tube formation assays, which revealed that siALKBH5 inhibited tube formation (Fig.6 and 6D), that produced the opposite effect by ALKBH5 overexpression (Fig.6 and 6F). Additionally, knockdown ALKBH5 in HUVECs downregulated the protein expression levels of p-AKT and p-mTOR in AKT/mTOR pathway, which were upregulated after ALKBH5 overexpression (Fig.6 and 6H). Overall, these data suggested that ALKBH5 played a role in regulating angiogenesis in endothelial cells, potentially through the AKT-mTOR signaling pathway.
4 Discussion
To explore the role of ALKBH5 in psoriasis angiogenesis, our study began by assessing ALKBH5 levels in both IMQ-treated and control mice. We used immunofluorescence co-staining to detect ALKBH5 and CD31 in the superficial dermis, noting elevated ALKBH5 in endothelial cells in IMQ-treated mice compared with the control group. This increase supports earlier findings of ALKBH5’s overexpression in various inflammatory skin conditions and cancers [
29,
30,
37]. In psoriasis, VEGF triggers endothelial cell differentiation and proliferation, enhancing angiogenesis [
38]. Our data suggest a crucial role for ALKBH5 in modulating angiogenesis, corroborated by both
in vivo and
in vitro experiments. Previous studies have also connected ALKBH5 overexpression in RPE cells with increased VEGFA secretion [
31,
33].
Further, administering varying doses of the ALKBH5 inhibitor IOX1 in an IMQ-induced PsD mouse model led to a significant reduction in psoriatic dermatitis, supported by PASI scores and histological reviews of reduced skin thickness and improved symptoms. Angiogenesis in the skin also diminished with IOX1 treatment, evidenced by reduced vascular development and lower VEGF protein expression. Higher doses of IOX1 intensified these therapeutic effects. Considering the inhibitory impact of IOX1 on ALKBH5, we hypothesize that the upregulation of ALKBH5 in psoriatic mice exacerbates psoriatic-like inflammation, angiogenesis, and epidermal proliferation as a compensatory mechanism. Similar to our results, IOX1 was found to effectively reduce IL-17 expression in CD4
+ T cells, inhibiting Th17 cell migration to inflammation sites [
39]. Therefore, we speculate that IOX1 might reduce the symptoms of psoriasis by inhibiting IL-17 in CD4
+ T cells. Utilizing ALKBH5-KO mice in IMQ models also showed reduced psoriatic dermatitis symptoms compared to WT counterparts. This was marked by decreased skin inflammation, thinner epidermis, and lower mRNA levels of CXCL12 and inflammatory cytokines like IL-17A and IL-1β in skin lesions. Additionally, skin angiogenesis significantly declined in ALKBH5-KO mice, with reductions in VEGF and VEGFR expression, decreased ALKBH5 and CD31 co-expression, and downregulated VEGFA, VEGFR2, p-AKT, and p-mTOR proteins in psoriatic mice. Our phenotypic experiments revealed that in a psoriasis-like mouse model, the knockout group showed reduced skin angiogenesis and epidermal thickness compared to the control group, suggesting a loss of target protein function. This aligns with previous studies showing that ALKBH5 knockout alleviates abnormal cell proliferation and vascular hyperplasia. We propose that the antibody’s specificity may leave protein residues post-knockout. Previous studies have indicated that suppressing or deleting ALKBH5 mitigates symptoms in melanoma and other inflammatory skin diseases effectively [
29,
30,
40].
Angiogenesis, involving the migration and proliferation of endothelial cells, is critical in psoriasis pathogenesis [
6]. We used IL-17A-induced HUVECs
in vitro to study angiogenesis [
7,
41], testing other inflammatory factors in HUVEC and HaCat cells. Notably, ALKBH5 protein and RNA levels significantly increased in IL-17A-induced HUVECs, while other settings showed no change. IL-17A treatment also reduced the m
6A ratio in HUVECs, suggesting a relationship between decreased m
6A levels in endothelial cells and IL-17A treatment [
42], supporting our previous findings on IL-17A-induced upregulation of ALKBH5 demethylase in HUVECs.
In vitro assays using CCK-8 and tube formation showed that ALKBH5 knockdown in HUVECs led to decreased cell proliferation and angiogenesis, along with lower IL-17RA mRNA and VEGFA, VEGFR2, p-AKT, and p-mTOR protein levels. Conversely, ALKBH5 overexpression increased these activities. These results suggest that ALKBH5 critically influences angiogenesis and inflammation in PsD through the AKT-mTOR pathway. Further studies are essential to explore ALKBH5’s role in psoriasis pathogenesis through this signaling pathway.
Our study identified increased ALKBH5 expression in psoriatic skin and its role in promoting angiogenesis. However, the exact mechanism by which IL-17A is downregulated following ALKBH5 deletion in psoriatic mice skin remains unclear, requiring further experimental investigation. Suppression of IL-17A in IMQ-induced γδT cells in psoriatic mice reduces inflammation [
43], while another study shows that deleting ALKBH5 boosts γδT cell precursor development by increasing m
6A levels [
44]. Thus, ALKBH5 may regulate γδT cell maturation and distribution in psoriasis, highlighting the need for more research to understand their interaction. Overall, ALKBH5 contributes to psoriasis progression and could be a promising therapeutic target.
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