Genetic perturbation of IFN-α transcriptional modulators in human endothelial cells uncovers pivotal regulators of angiogenesis

Ciccarese, Francesco; Grassi, Angela; Pasqualini, Lorenza; Rosano, Stefania; Noghero, Alessio; Montenegro, Francesca; Bussolino, Federico; Camillo, Barbara Di; Finesso, Lorenzo; Toffolo, Gianna; Mitola, Stefania; Indraccolo, Stefano

doi:10.1016/j.csbj.2020.11.048

Cited by 7 publications

(3 citation statements)

References 53 publications

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“…Indeed, ectopic expression of IFIT1 and IFIT3 has been shown to enhance many EGFR and annexin-2 pathways to induce cell proliferation, survival, and drug resistance [54]. Additionally, we observed downregulation of USP18, which mediates angiogenic effects of IFN-α by promoting EC capillary sprouting and angiogenesis [92,93]. Moreover, USP18 deficiency in mammary cells creates antitumor activity [94]; L-GALS9 which induces angiogenesis and MAPK phosphorylation in VECs [95] and galectins play a critical role in regulating angiogenesis [96].…”

Section: Discussionmentioning

confidence: 71%

Micro-RNA193a-3p Inhibits Breast Cancer Cell Driven Growth of Vascular Endothelial Cells by Altering Secretome and Inhibiting Mitogenesis: Transcriptomic and Functional Evidence

Azzarito

Kurmann

Leeners

et al. 2022

Cells

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Breast cancer (BC) cell secretome in the tumor microenvironment (TME) facilitates neo-angiogenesis by promoting vascular endothelial cell (VEC) growth. Drugs that block BC cell growth or angiogenesis can restrict tumor growth and are of clinical relevance. Molecules that can target both BC cell and VEC growth as well as BC secretome may be more effective in treating BC. Since small non-coding microRNAs (miRs) regulate cell growth and miR193a-3p has onco-suppressor activity, we investigated whether miR193a-3p inhibits MCF-7-driven growth (proliferation, migration, capillary formation, signal transduction) of VECs. Using BC cells and VECs grown in monolayers or 3D spheroids and gene microarrays, we demonstrate that: pro-growth effects of MCF-7 and MDA-MB231 conditioned medium (CM) are lost in CM collected from MCF-7/MDA-MB231 cells pre-transfected with miR193a-3p (miR193a-CM). Moreover, miR193a-CM inhibited MAPK and Akt phosphorylation in VECs. In microarray gene expression studies, miR193a-CM upregulated 553 genes and downregulated 543 genes in VECs. Transcriptomic and pathway enrichment analysis of differentially regulated genes revealed downregulation of interferon-associated genes and pathways that induce angiogenesis and BC/tumor growth. An angiogenesis proteome array confirmed the downregulation of 20 pro-angiogenesis proteins by miR193a-CM in VECs. Additionally, in MCF-7 cells and VECs, estradiol (E2) downregulated miR193a-3p expression and induced growth. Ectopic expression of miR193a-3p abrogated the growth stimulatory effects of estradiol E2 and serum in MCF-7 cells and VECs, as well as in MCF-7 and MCF-7+VEC 3D spheroids. Immunostaining of MCF-7+VEC spheroid sections with ki67 showed miR193a-3p inhibits cell proliferation. Taken together, our findings provide first evidence that miR193a-3p abrogates MCF-7-driven growth of VECs by altering MCF-7 secretome and downregulating pro-growth interferon signals and proangiogenic proteins. Additionally, miR193a-3p inhibits serum and E2-induced growth of MCF-7, VECs, and MCF-7+VEC spheroids. In conclusion, miRNA193a-3p can potentially target/inhibit BC tumor angiogenesis via a dual mechanism: (1) altering proangiogenic BC secretome/TME and (2) inhibiting VEC growth. It may represent a therapeutic molecule to target breast tumor growth.

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Section: Discussionmentioning

confidence: 71%

Micro-RNA193a-3p Inhibits Breast Cancer Cell Driven Growth of Vascular Endothelial Cells by Altering Secretome and Inhibiting Mitogenesis: Transcriptomic and Functional Evidence

Azzarito

Kurmann

Leeners

et al. 2022

Cells

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“…Interestingly, ectopic expression of IFIT1 and IFIT3 enhances many EGFR and annexin-2 pathways to induce cell proliferation, survival, and drug resistance [71]. Moreover, USP18, which mediates angiogenic effects (sprouting and capillary formation) of IFN-α in ECs [96,97], was downregulated by miR193a-3p in spheroids. The above findings, together with the fact that USP18 deficiency creates antitumor activity in mammary cells [98], suggest that miR193a-3p may prevent MCF-7 and LEC cross-talk and abrogate IFN-driven angiogenic processes in BC, inhibiting growth.…”

Section: Discussionmentioning

confidence: 99%

Transcriptomic and Functional Evidence That miRNA193a-3p Inhibits Lymphatic Endothelial Cell (LEC) and LEC + MCF-7 Spheroid Growth Directly and by Altering MCF-7 Secretome

Azzarito

Henry

Rotshteyn

et al. 2023

Cells

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MicroRNA 193a-3p (miR193a-3p) is a short non-coding RNA with tumor suppressor properties. Breast cancer (BC) progression is governed by active interaction between breast cancer cells, vascular (V)/lymphatic (L) endothelial cells (ECs), and BC secretome. We have recently shown that miR193a-3p, a tumor suppressor miRNA, inhibits MCF-7 BC cell-driven growth of VECs via direct antimitogenic actions and alters MCF-7 secretome. Since LEC-BC cross-talk plays a key role in BC progression, we investigated the effects of miR193a-3p on MCF-7 secretome and estradiol-mediated growth effects in LECs and LEC + MCF-7 spheroids, and delineated the underlying mechanisms. Transfection of LECs with miR193a-3p, as well as secretome from MCF-7 transfected cells, inhibited LEC growth, and these effects were mimicked in LEC + MCF-7 spheroids. Moreover, miR193a-3p inhibited ERK1/2 and Akt phosphorylation in LECs and LEC + MCF-7 spheroids, which are importantly involved in promoting cancer development and metastasis. Treatment of LECs and LEC + MCF-7 spheroids with estradiol (E2)-induced growth, as well as ERK1/2 and Akt phosphorylation, and was abrogated by miR193a-3p and secretome from MCF-7 transfected cells. Gene expression analysis (GEA) in LEC + MCF-7 spheroids transfected with miR193a-3p showed significant upregulation of 54 genes and downregulation of 73 genes. Pathway enrichment analysis of regulated genes showed significant modulation of several pathways, including interferon, interleukin/cytokine-mediated signaling, innate immune system, ERK1/2 cascade, apoptosis, and estrogen receptor signaling. Transcriptomic analysis showed downregulation in interferon and anti-apoptotic and pro-growth molecules, such as IFI6, IFIT1, OSA1/2, IFITM1, HLA-A/B, PSMB8/9, and PARP9, which are known to regulate BC progression. The cytokine proteome array of miR193a-3p transfected MCF secretome and confirmed the upregulation of several growth inhibitory cytokines, including IFNγ, Il-1a, IL-1ra, IL-32, IL-33, IL-24, IL-27, cystatin, C-reactive protein, Fas ligand, MIG, and sTIM3. Moreover, miR193a-3p alters factors in MCF-7 secretome, which represses ERK1/2 and Akt phosphorylation, induces pro-apoptotic protein and apoptosis in LECs, and downregulates interferon-associated proteins known to promote cancer growth and metastasis. In conclusion, miR193a-3p can potentially modify the tumor microenvironment by altering pro-growth BC secretome and inhibiting LEC growth, and may represent a therapeutic molecule to target breast tumors/cancer.

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“…ZFPM2-AS1 has been shown to play crucial roles in the progression of retinoblastoma, an angiogenesis-associated disease [ 45 ]. The interferon (IFN) family of cytokines serves as pivotal regulators of angiogenesis [ 46 ], while NRIR was found to be a negative Regulator of IFN Response [ 47 , 48 ]. Tumor growth of Clear Cell Renal Cell Carcinoma was suppressed by silenced LINC01215 via miR-184 [ 49 ].…”

Section: Discussionmentioning

confidence: 99%

Galectin-1-dependent ceRNA network in HRMECs revealed its association with retinal neovascularization

et al. 2023

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Background Retinal neovascularization (RNV) is a leading cause of blindness worldwide. Long non-coding RNA (lncRNA) and competing endogenous RNA (ceRNA) regulatory networks play vital roles in angiogenesis. The RNA-binding protein galectin-1 (Gal-1) participates in pathological RNV in oxygen-induced retinopathy mouse models. However, the molecular associations between Gal-1 and lncRNAs remain unclear. Herein, we aimed to explore the potential mechanism of action of Gal-1 as an RNA-binding protein. Results A comprehensive network of Gal-1, ceRNAs, and neovascularization-related genes was constructed based on transcriptome chip data and bioinformatics analysis of human retinal microvascular endothelial cells (HRMECs). We also conducted functional enrichment and pathway enrichment analyses. Fourteen lncRNAs, twenty-nine miRNAs, and eleven differentially expressed angiogenic genes were included in the Gal-1/ceRNA network. Additionally, the expression of six lncRNAs and eleven differentially expressed angiogenic genes were validated by qPCR in HRMECs with or without siLGALS1. Several hub genes, such as NRIR, ZFPM2-AS1, LINC0121, apelin, claudin-5, and C-X-C motif chemokine ligand 10, were found to potentially interact with Gal-1 via the ceRNA axis. Furthermore, Gal-1 may be involved in regulating biological processes related to chemotaxis, chemokine-mediated signaling, the immune response, and the inflammatory response. Conclusions The Gal-1/ceRNA axis identified in this study may play a vital role in RNV. This study provides a foundation for the continued exploration of therapeutic targets and biomarkers associated with RNV.

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Genetic perturbation of IFN-α transcriptional modulators in human endothelial cells uncovers pivotal regulators of angiogenesis

Abstract: Graphical abstract

Cited by 7 publications

References 53 publications

Micro-RNA193a-3p Inhibits Breast Cancer Cell Driven Growth of Vascular Endothelial Cells by Altering Secretome and Inhibiting Mitogenesis: Transcriptomic and Functional Evidence

Micro-RNA193a-3p Inhibits Breast Cancer Cell Driven Growth of Vascular Endothelial Cells by Altering Secretome and Inhibiting Mitogenesis: Transcriptomic and Functional Evidence

Transcriptomic and Functional Evidence That miRNA193a-3p Inhibits Lymphatic Endothelial Cell (LEC) and LEC + MCF-7 Spheroid Growth Directly and by Altering MCF-7 Secretome

Galectin-1-dependent ceRNA network in HRMECs revealed its association with retinal neovascularization

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