Tao Zhang,Qianhan Yang,Jun Chen,Jiyu Jiang,Jingnan Han,Yun Zhou
Diabetic retinopathy (DR), the leading cause of adult blindness, has its risk increased by excessive endoplasmic reticulum stress (ERS). Nuclear receptor subfamily 2 group f member 2 (NR2F2) is an orphan nuclear receptor with essential roles in angiogenesis. However, its roles in DR remain unknown. Notably, NR2F2 protein expression was upregulated in streptozotocin-induced diabetic mice retina and the retinal endothelial cells characterized by endothelial marker CD31. Retinal NR2F2 expression was knocked down in diabetic mice using adeno-associated virus serotype 2. NR2F2 knockdown increased retinal thickness, decreased acellular capillary formation and FITC-labeled dextran leakage, and increased tight junction (TJ) protein ZO-1 expression in vivo. Additionally, NR2F2 knockdown inhibited reactive oxygen species generation and the ERS marker CHOP expression in vivo. In high glucose (HG)-induced human retinal microvascular endothelial cell (HRMEC) monolayers, NR2F2 knockdown inhibited FITC-dextran flux, increased transendothelial electrical resistance, reduced VEGFA secretion and restored expression and continuous distribution of TJ proteins. NR2F2 knockdown also reduced ERS markers expression and inhibited ERS-related signaling pathways in vitro. mRNA-seq analysis in NR2F2-knockdown HRMECs exposed to HG showed that NR2F2 inhibition upregulated CYB5R2 mRNA level. NR2F2 was bound directly to the CYB5R2 promoter for transcriptional repression in HRMECs. CYB5R2 overexpression inhibited ERS-related protein expressions and barrier dysfunction in HRMEC monolayers, and CYB5R2 knockdown reversed the endothelial protective effect of NR2F2 inhibition. In conclusion, NR2F2 knockdown attenuates vascular dysfunction by alleviating ERS in retinal microvascular endothelial cells and alleviates disease progression in DR mice. This may be achieved by reducing CYB5R2 transcriptional repression. Graphical Nuclear receptor subfamily 2 group f member 2 (NR2F2) knockdown increased the NADH-Cytochrome b5 reductase 2 (CYB5R2) expression in human retinal microvascular endothelial cells by reducing CYB5R2 transcriptional repression. The upregulation of CYB5R2 attenuated vascular dysfunction by inhibiting endoplasmic reticulum stress (ER Stress)-related pathways including PERK, IRE1, and ATF6 pathways in retinal microvascular endothelial cells, thereby inhibiting disease progression in diabetic retinopathy mice.
Purpose This study aims to investigate the role of AGAP2 in acute myeloid leukemia (AML), particularly its involvement in lipid metabolic reprogramming and AML progression, and to explore the underlying regulatory mechanisms. Methods AGAP2 expression was analyzed in CD34⁺ cells from AML patients stratified by cytogenetic risk. Functional assays, including knockdown and overexpression of AGAP2, were performed in KG-1 and HL-60 AML cell lines to assess effects on proliferation, apoptosis, differentiation, and lipid metabolism. mRNA sequencing and KEGG pathway analysis were used to identify relevant signaling pathways. Lipid accumulation was evaluated via Filipin III staining. The AMPK pathway activator GSK621 was applied to examine lipid deposition. In vivo tumor growth and survival were studied in a mouse model, and bone marrow infiltration of CD45⁺ cells was analyzed. The transcriptional regulation of AGAP2 by NR2F2 was assessed through promoter binding and functional rescue experiments. Results AGAP2 was highly expressed in CD34⁺ cells from AML patients, with higher levels in the poor-risk group compared to favorable- and intermediate-risk groups. Knockdown of AGAP2 induced apoptosis and differentiation, while its overexpression promoted cell proliferation. KEGG analysis indicated enrichment of differentially expressed genes in the AMPK pathway. AGAP2 was found to inhibit AMPK/ACC pathway activation. Overexpression of AGAP2 increased lipid deposition whereas GSK621 treatment reversed it. In vivo, AGAP2 knockdown suppressed tumor growth, improved survival, and reduced CD45⁺ cell infiltration in bone marrow. Furthermore, NR2F2 bound to the AGAP2 promoter and suppressed its transcription. NR2F2 overexpression reduced lipid accumulation, an effect reversed by AGAP2 overexpression. Conclusion AGAP2 promotes lipid deposition and AML progression by inhibiting the AMPKα/ACC pathway. NR2F2 acts as a transcriptional repressor of AGAP2, thereby attenuating AML development. These findings reveal a novel NR2F2/AGAP2/AMPK/ACC regulatory axis in AML, highlighting potential therapeutic targets for lipid metabolism-based interventions in leukemia. Graphical
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