JB and VL are recipients of Jnos Bolyai Research Scholarship of the Hungarian Academy of Sciences, VL is supported by UNKP-19-4 New National Superiority Program of the Ministry for Development and Technology, and ZM by UNKP-20-3.?? We also thank Berta Kamprad Foundation, The Mats Paulsson Trust and The Stefan Paulsson Trust for their kind support. Data availability The datasets used and analyzed during the current study are available from your corresponding authors on reasonable request. Competing interests The authors declare no competing interests. Footnotes Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations. These authors contributed equally: Balzs D?me and Viktria Lszl. Contributor Information Balzs D?me, Email: firstname.lastname@example.org. Viktria Lszl, Email: email@example.com. Supplementary Information The online version contains supplementary material available at 10.1038/s41598-021-85162-0.. in tumor angiogenesis and growth in various experimental systems. We investigated the role of apelin signaling in the malignant behavior of cutaneous melanoma. Murine B16 and human A375 melanoma cell lines were stably transfected with apelin encoding or control vectors. Apelin overexpression significantly increased melanoma cell migration and invasion in vitro, but it experienced no impact on its proliferation. In our in vivo experiments, apelin significantly increased the number and size of lung metastases of murine melanoma cells. Melanoma cell proliferation rates and lymph and blood microvessel densities were significantly higher in the apelin-overexpressing pulmonary metastases. APJ inhibition by the competitive APJ antagonist MM54 significantly attenuated the in vivo pro-tumorigenic Schisanhenol effects of apelin. Additionally, we detected significantly elevated circulating apelin and VEGF levels in patients with melanoma compared to healthy controls. Our results show that apelin promotes blood and lymphatic vascularization and the growth of pulmonary metastases of skin melanoma. Further studies are warranted to validate apelin signaling as a new potential therapeutic target in this malignancy. show the colony of the B16 Mock (A) or B16 Ap (B,C) melanoma cells in the lung. (D,E) Apelin overexpression significantly enhanced the number (D) and area (E) of B16 Ap lung metastases compared to Schisanhenol the metastases of B16 Mock cells. Furthermore, the APJ antagonist MM54 experienced no impact on the number of metastases (D), but it significantly decreased the area of Schisanhenol lung metastases created by B16 Ap cells (E). *superficial malignant melanoma, nodular malignant melanoma, acral lentiginous melanoma, not available. value?=?0.0405; Supp Fig.?3B)55. In addition, Tumor Immune Estimation Resource 2.0 (TIMER 2.0) and the Human Protein Atlas also showed a tendency towards poor prognosis in patients with high apelin levels, when analyzing the associations between the expression levels of apelin gene and survival outcomes in TCGA: TIMER 2.0 analyzed 471 patients with skin cutaneous melanoma (test if the sample distribution was normal or with MannCWhitney U test if the sample distribution was asymmetric. Tukeys multiple comparison test were utilized for more than two groups. Differences were decided to be significant if p?0.05. All statistical analyses were carried out using GraphPad Prism 6.0 (GraphPad Software, Inc., La Jolla, CA, USA) and IBM SPSS Statistics 23 software (IBM Corp., Armonk, NY, USA). Ethics approval and consent to participate The study was conducted in accordance with the current National Comprehensive Malignancy Network guidelines, based on the ethical requirements prescribed by the Helsinki Declaration of the World Medical Association. All patients and controls experienced given informed consent, and the sample collection was approved by the Regional Ethical Review Table in Lund (Approval Figures: DNR 2013/101 and DNR 2013/480). The animals used in this study were cared according to the Guiding Principles for the Care and Use of Animals based upon the Helsinki declaration. The animal-model experiments were carried Ankrd1 out in compliance with the ARRIVE guidelines and approved by the Ethics Committee of the Department of Food Chain Safety, Animal Health, Plant and Soil Protection, Pest County Government Office (License Number: PEI/001/2574-6/2015). Supplementary Information Supplementary Video 1.(54M, avi) Supplementary Video 2.(41M, avi) Supplementary Video 3.(5.3M, avi) Supplementary Video 4.(5.6M, avi) Supplementary Information 1.(1.8M, tif) Supplementary Information 2.(566K, tif) Supplementary Information 3.(645K, tif) Supplementary Information 4.(86K, pdf) Acknowledgements We would like to thank Dra Lakatos, the member of the Department of Biological Physics, E?tv?s University or college for helping to evaluate the videomicroscopic measurements. We also specially thank Zsfia Szab-Zsibai for scanning the microscope slides, and Katalin Parragn Derecskei and Barbara Dekan for their excellent technical assistance. Abbreviations MEKMitogen-activated protein kinase kinaseVEGFVascular endothelial growth factorRT-qPCRReverse transcription-quantitative polymerase chain reactionECMExtracellular matrixBrdU5-Bromo-2-deoxyuridineGEPIA2Gene expression profiling interactive analysis 2TIMER 2.0Tumor immune estimation resource 2.0SRBSulforhodamine BErkExtracellular signal-regulated kinaseMMP-1Matrix metalloproteinase-1MMP-9Matrix metalloproteinase-9IL-2Interleukin-2IL-6Interleukin-6EDTAEthylenediaminetetraacetic acidELISAEnzyme-linked immunosorbent assayDMEMDulbeccos modified Eagles mediumFBSFetal bovine serumPIVParticle image velocimetryPBSPhosphate buffered salineIHCImmunohistochemical stainingBSABovine serum albuminLYVE-1Lymphatic vessel endothelial receptor 1 Author contributions J.B. performed the experiments, analyzed the data and published the paper. S.T., O.D. and I.K. contributed Schisanhenol to the in vivo experiments. J.T.Z. performed and analyzed the videomicroscopic measurements. H.O. participated in collection of.