BACKGROUND: Psoriasis is an immune-mediated chronic inflammatory skin disease. Current research suggests that the long-term persistence and recurrence of psoriasis are closely related to the feedback loop formed between keratinocytes and immune cells, especially in Th 17 or DC cells expressing CCR6. CCL20 is the ligand of CCR6. Therefore, drugs that block the expression of CCL20 or CCR6 may have a certain therapeutic effect on psoriasis. Glycyrrhetinic acid (GA) is the main active ingredient of the plant drug licorice and is often used to treat autoimmune diseases, including psoriasis. However, its mechanism of action is still unclear.
METHODS: Psoriasis like skin lesion model was established by continuously applying imiquimod on the back skin of normal mice and CCR6-/- mice for 7 days. The therapeutic and preventive effects of glycyrrhetinic acid (GA) on the model were observed and compared. The severity of skin injury is estimated through clinical PASI scores and histopathological examination. qRT-PCR and multiple cytoline assay were explored to detect the expression levels of cytokines in animal dorsal skin lesions and keratinocyte line HaCaT cells, respectively. The dermis and epidermis of the mouse back were separated for the detection of CCL20 expression. Transcription factor assay was applied to screen, and luciferase activity assay to validate transcription factors regulated by GA. Technology of surface plasmon laser resonance with LC-MS (SPR-MS), molecular docking, and enzyme activity assay were used to identified the target proteins for GA. Finally, we synthesized different derivatives of 18beta-GA and compared their effects, as well as glycyrrhetinic acid (GL), on the skin lesion of imiquimod-induced mice to evaluate the active groups of 18beta-GA.
RESULTS: 18β-glycyrrhetinic acid (GA) improved IMQ-induced psoriatic lesions, and could specifically reduce the chemokine CCL20 level of the epidermis in lesion area, especially in therapeutic administration manner. The process was mainly regulated by transcription factor ATF2 in the keratinocytes. In addition, GUSB was identified as the primary target of 18βGA. Our findings indicated that the subject on molecular target research of glycyrrhizin should be glycyrrhetinic acid (GA) instead of glycyrrhizic acid (GL), because GL showed little activity in vitro or in vivo. Apart from that, α, β, -unsaturated carbonyl in C11/12 positions was crucial or unchangeable to its activity of 18βGA, while proper modification of C3 or C30 position of 18βGA may vastly increase its activity.
CONCLUSIONS: Our research indicates that 18βGA exerted its anti-psoriasis effect mainly by suppressing ATF2 and downstream molecule CCL20 predominately through α, β, -unsaturated carbonyl at C11/12 position binding to GUSB in the keratinocytes, and then broke the feedback loop between keratinocytes and CCR6-expressing immune cells. GA has more advantages than GL in the external treatment of psoriasis. A highlight of this study is to investigate the influence of special active groups on the pharmacological action of a natural product, inspired by the molecular docking result.

BACKGROUND: Ankyrin repeat domain 49 (ANKRD49) has been found to be highly expressed in multiple cancer including lung adenocarcinoma (LUAD) and lung squamous carcinoma (LUSC). However, the function of ANKRD49 in the pathogenesis of NSCLC still remains elusive. Previously, ANKRD49 has been demonstrated to promote the invasion and metastasis of A549 cells, a LUAD cell line, via activating the p38-ATF-2-MMP2/MMP9 pathways. Considering the heterogeneity of tumor cells, the function and mechanism of ANKRD49 in NSCLC need more NSCLC-originated cells to clarify.
METHODS: Real-time qPCR was employed to test ANKRD49 expression levels in nine pairs of fresh NSCLC tissues and the corresponding adjacent normal tissues. The function of ANKRD49 was investigated using overexpression and RNA interference assays in lung adenocarcinoma cell line (NCI-H1299) and lung squamous carcinoma cell line (NCI-H1703) through gelatin zymography, cell counting kit-8, colony formation, wound healing, migration and invasion assays mmunoprecipitation was performed to in vitro. Immunoprecipitation was performed to test the interaction of c-Jun and ATF2. Chromatin immunoprecipitation was conducted to assess the transcriptional regulation of ATF2/c-Jun on MMP-2/9. Moreover, the tumorigenicity of ANKRD49 was evaluated in nude mice models and the involved signal molecular was also measured by immunohistochemical method.
RESULTS: We found that the levels of ANKRD49 in cancerous tissues were higher than those in adjacent normal tissues. in vitro assay showed that ANKRD49 promoted the migration and invasion of NCI-H1299 and NCI-H1703 cells via enhancing the levels of MMP-2 and MMP-9. Furthermore, ANKRD49 elevated phosphorylation of JNK and then activated c-Jun and ATF2 which interact in nucleus to promote the binding of ATF2:c-Jun with the promoter MMP-2 or MMP-9. In vivo assay showed that ANKRD49 promoted lung metastasis of injected-NSCLC cells and the high metastatic rate was positively correlated with the high expression of ANKRD49, MMP-2, MMP-9, p-JNK, p-c-Jun and p-ATF2.
CONCLUSIONS: The present study indicated that ANKRD49 accelerated the invasion and metastasis of NSCLC cells via JNK-mediated transcription activation of c-Jun and ATF2 which regulated the expression of MMP-2/MMP-9. The molecular mechanisms of ANKRD49\'s function is different from those found in A549 cells. The current study is a supplement and improvement to the previous research.

Activating transcription factor 2 (ATF2) is a member of the leucine zipper family of DNA binding proteins that are responsible for regulating various genes that play an essential role in major biological and cellular functions. Since ATF2 plays a vital role in cellular proliferation and apoptosis, it is believed that it greatly affects the development of breast cancers. However, its exact role in breast cancer is incompletely understood. It remains a subject of debate, ambiguity, and continuous research. Several studies have suggested the role of ATF2 as an oncogene, promoting cellular proliferation and worsening the outcome of cancers. In contrast, other studies have postulated that ATF2 plays a tumor suppressive role in estrogen receptor-positive breast cancer. The ambiguity surrounding its role in breast cancer is the reason why there is an influx of recent studies and research in this area. In this narrative review, we investigate several studies that have been published about the role of ATF2 in breast cancer. We also explore studies that have examined the association between ATF2 and endocrine therapy resistance. ATF2 has been suggested to modulate estrogen receptor (ER) expression and activity, potentially affecting tamoxifen sensitivity in breast cancer cells. Therefore, the role of ATF2 in DNA repair mechanisms and drug resistance has been deeply explored in this review. Additionally, there are numerous ongoing clinical trials exploring the effect of targeting ATF2 pathways and mechanisms on the outcome of breast cancers, some of which we have discussed. The studies and clinical trials that are being conducted to understand the multifaceted role of ATF2 and its signaling pathways may provide valuable insight for developing efficient targeted therapeutic solutions to enhance the outcomes of breast cancer and overcome endocrine resistance. We suggest further research to elucidate the dual roles of ATF2 in breast cancer and potential therapeutic therapies for its treatment.

BACKGROUND: Polymethylmethacrylate (PMMA) bone cement loaded with enoxaparin sodium (PMMA@ES) has been increasingly highlighted to affect the bone repair of bone defects, but the molecular mechanisms remain unclear. We addressed this issue by identifying possible molecular mechanisms of PMMA@ES involved in femoral defect regeneration based on bioinformatics analysis and network pharmacology analysis.
METHODS: The upregulated genes affecting the osteogenic differentiation of bone marrow mesenchymal stem cells (BMSCs) were selected through bioinformatics analysis, followed by intersection with the genes of ES-induced differentiation of BMSCs identified by network pharmacology analysis. PMMA@ES was constructed. Rat primary BMSCs were isolated and cultured in vitro in the proliferation medium (PM) and osteogenic medium (OM) to measure alkaline phosphatase (ALP) activity, mineralization of the extracellular matrix, and the expression of RUNX2 and OCN using gain- or loss-of-function experiments. A rat femoral bone defect model was constructed to detect the new bone formation in rats.
RESULTS: ATF2 may be a key gene in differentiating BMSCs into osteoblasts. In vitro cell assays showed that PMMA@ES promoted the osteogenic differentiation of BMSCs by increasing ALP activity, extracellular matrix mineralization, and RUNX2 and OCN expression in PM and OM. In addition, ATF2 activated the transcription of miR-335-5p to target ERK1/2 and downregulate the expression of ERK1/2. PMMA@ES induced femoral defect regeneration and the repair of femoral defects in rats by regulating the ATF2/miR-335-5p/ERK1/2 axis.
CONCLUSIONS: The evidence provided by our study highlighted the ATF2-mediated mechanism of PMMA@ES in the facilitation of the osteogenic differentiation of BMSCs and femoral defect regeneration.

Amentoflavone (AF) is a natural multifunctional biflavonoid that has been revealed to possess multiple biological activities, including anticancer activity. Here, this work focused on exploring the functions and mechanism of AF in gastric cancer (GC). Levels of genes and proteins were examined by quantitative real-time PCR and western blotting. Cell proliferation and cell death were analyzed using cell counting kit-8, colony formation, and lactate dehydrogenase (LDH) release assay, respectively. Cell ferroptosis was evaluated by detecting the levels of malondialdehyde (MDA), reduced glutathione (GSH), Fe2+ , and intracellular reactive oxygen species (ROS). The binding between miR-496 and activating transcription factor 2 (ATF2) was confirmed by using dual-luciferase reporter assay. Murine xenograft assay was conducted for in vivo experiments. The results showed that AF suppressed the proliferation and induced ferroptotic cell death in GC cells. MiR-496 expression was decreased in GC tissues and cells, and AF treatment increased miR-496 expression level in GC cells. Functionally, miR-496 inhibition reversed the inhibitory effects of AF on GC cell proliferation and promoting effects on ferroptotic cell death. Mechanistically, ATF2 was targeted by miR-496. ATF2 expression was increased in GC tissues and cells, which was decreased by AF treatment and subsequently rescued by miR-496 downregulation in GC cells. Moreover, miR-496 overexpression suppressed the proliferation and induced ferroptotic cell death in GC cells via targeting ATF2. In all, AF suppressed the proliferation and induced ferroptotic cell death in GC cells via miR-496/ATF2 axis, indicating a novel therapeutic approach for GC patients.

Globally, gastric cancer (GC) is a major cause of cancer death. This study is aimed at investigating the biological functions of activating transcription factor 2 (ATF2) and the underlying mechanism in GC. In the present work, GEPIA, UALCAN, Human Protein Atlas and StarBase databases were adopted to analyze ATF2 expression characteristics in GC tissues and normal gastric tissues, and its relationships with tumor grade and patients\' survival time. Quantitative real-time polymerase chain reaction (qRT-PCR) method was employed to examine ATF2 mRNA expression in normal gastric tissues, GC tissues, and GC cell lines. Cell counting kit-8 (CCK-8) and EdU assays were utilized for detecting GC cell proliferation. Cell apoptosis was detected by flow cytometry. PROMO database was applied to predict the binding site of ATF2 with the METTL3 promoter region. The binding relationship between ATF2 and the METTL3 promoter region was verified through dual-luciferase reporter gene assay and chromatin immunoprecipitation-quantitative PCR (ChIP-qPCR) assay. Western blot was performed to evaluate the effect of ATF2 on METTL3 expression. METTL3-related signaling pathways were predicted using Gene Set Enrichment Analysis (GSEA) in the LinkedOmics database. It was found that, ATF2 level was elevated in GC tissues and cell lines in comparison with normal tissues and correlated with short patients\' survival time. ATF2 overexpression facilitated GC cell growth and suppressed the apoptosis, whereas ATF2 knockdown suppressed GC cell proliferation and facilitated the apoptosis. ATF2 bound to the METTL3 promoter region, and ATF2 overexpression promoted the transcription of METTL3, and ATF2 knockdown restrained the transcription of METTL3. METTL3 was associated with cell cycle progression, and ATF2 overexpression enhanced cyclin D1 expression, and METTL3 knockdown reduced cyclin D1 expression. In summary, ATF2 facilitates GC cell proliferation and suppresses the apoptosis via activating the METTL3/cyclin D1 signaling pathway, and ATF2 is promising to be an anti-drug target for GC.

The differentiation and activation of macrophages are critical regulatory programs that are central to host inflammation and pathogen defense. However, the transcriptional regulatory pathways involved in these programs are not well understood. Herein, we demonstrate that the activity and expression of the transcription factor ATF2 is precisely regulated during primary human monocyte-to-macrophage differentiation and that its activation is linked to M1 polarization and antibacterial responses. Genetic perturbation experiments demonstrated that deletion of ATF2 (THP-ΔATF2) resulted in irregular and abnormal macrophage morphology, whereas macrophages overexpressing ATF2 (THP-ATF2) developed round and pancake-like morphology, resembling classically activated (M1) macrophages. Mechanistically, we show that ATF2 binds to the core promoter of PPM1A, a phosphatase that regulates monocyte-to-macrophage differentiation, to regulate its expression. Functionally, overexpression of ATF2 sensitized macrophages to M1 polarization, resulting in increased production of major histocompatibility complex class II, IL-1β, and IP-10; improved phagocytic capacity; and enhanced control of the intracellular pathogen Mycobacterium tuberculosis. Gene expression profiling revealed that overexpression of ATF2 reprogramed macrophages to promote antibacterial pathways enriched in chemokine signaling, metabolism, and antigen presentation. Consistent with pathways analysis, metabolic profiling revealed that genetic overexpression or stimuli-induced activation of ATF2 alters the metabolic capacity of macrophages and primes these cells for glycolytic metabolism during M1 polarization or bacterial infection. Our findings reveal that ATF2 plays a central role during macrophage differentiation and M1 polarization to enhance the functional capacities of macrophages.

Diabetic retinopathy (DR) is one of the leading causes of blindness in diabetic patients. However, the pathogenesis of DR is complex, and no firm conclusions have been drawn so far. It has become a hot spot in ophthalmology research to deeply study the mechanism of DR pathological changes and find effective treatment options. Human retinal microvascular endothelial cells (HRMECs) were induced by high glucose (HG) to construct DR cell model. CCK-8 assay was used to detect the viability of HRMECs. Transwell assay was used to detect the migration ability of HRMECs. Tube formation assay was used to identify the tube formation ability of HRMECs. The expressions of USP14, ATF2 and PIK3CD were detected by Western blot analysis and qRT-PCR assay. Immunoprecipitation (IP) was used to ascertain the relationship of USP14 and ATF2. To explore the regulatory relationship between ATF2 and PIK3CD by dual-luciferase reporter gene assay and Chromatin immunoprecipitation (ChIP) assay. High glucose treatment promoted the proliferation, migration, and tube formation of HRMEC, and the expressions of USP14, ATF2 and PIK3CD were significantly up-regulated. USP14 or ATF2 knockdown inhibited HG-induced HRMECs proliferation, migration, and tube formation. USP14 regulated the expression of ATF2, and ATF2 promoted PIK3CD expression. PIK3CD overexpression attenuated the inhibitory effectiveness of USP14 knockdown on proliferation, migration and tube formation of DR cell model. Here, we revealed that USP14 regulated the ATF2/PIK3CD axis to promote proliferation, migration, and tube formation in HG-induced HRMECs.

Sorafenib, a tyrosine kinase inhibitor, has an important antitumor effect as a ferroptosis inducer in multiple cancers, including gastric cancer (GC). However, the status of sorafenib as a ferroptosis inducer has recently been questioned. There is very limited information about the relationship between ferroptosis and ATF2, and the role of ATF2 in sorafenib-induced ferroptosis has not been studied. In this study, we investigated the role and underlying molecular mechanisms of ATF2 in sorafenib-induced ferroptosis in GC. We found that ATF2 was significantly upregulated in GC tissues and predicted a poor clinical prognosis. Silencing ATF2 significantly inhibited the malignant phenotype of GC cells. In addition, we observed that ATF2 was activated during sorafenib-induced ferroptosis in GC cells. ATF2 knockdown promoted sorafenib-induced ferroptosis, while ATF2 overexpression showed the opposite results in GC cells. Using ChIP-Seq and RNA-Seq, we identified HSPH1 as a target of ATF2 and further validated it by ChIP‒qPCR analysis. HSPH1 can interact with SLC7A11 (cystine/glutamate transporter) and increase its protein stability. Importantly, knockdown of HSPH1 partly reversed the effects caused by ATF2 overexpression on sorafenib-induced ferroptosis in GC cells. In addition, the results from the tumor xenograft model showed that ATF2 knockdown can effectively enhance sorafenib sensitivity in vivo. Collectively, our study reveals a novel mechanism by which sorafenib induces ferroptosis in GC.

OBJECTIVE: To explore the ATF2 expression of preeclampsia patients and investigate whether the level of ATF2 expression impacted the low-dose aspirin treatment of preeclampsia patients.
BACKGROUND: Preeclampsia is a severe pregnancy-related hypertension disorder and refers to hypertension.
OBJECTIVE: This study was designed to explore the activating transcription factor 2 (ATF2) expression of preeclampsia patients and investigate whether the level of ATF2 expression impacted the low-dose aspirin treatment of preeclampsia patients.
METHODS: Firstly, we collected the plasma of normal and preeclampsia pregnancies and quantified the expressions of ATF2 by ELISA. Then we quantified the expression of the three downstream target genes of ATF2 (IL-8, IL-6 and MMP-2). Finally, we collected and quantified the interventional and observational group plasma. All data were compared by t-test (p<0.05).
RESULTS: ATF2 and its target genes (IL-6, IL-8 and MMP-2) were upregulated in preeclampsia patients. In addition, ATF2 and its target genes were downregulated in the interventional group (LDA-treated group).
CONCLUSIONS: Our results indicated that LDA could inhibit ATF2 expression in preeclampsia. It suggests that ATF2 may be a potential target of LDA in the prevention of preeclampsia.