Notch signaling controls sprouting angiogenesis of endometriotic lesions

Angiogenesis is essential for the engraftment and growth of endometriotic lesions. In this study, we analyzed whether this process is regulated by Notch signaling. Endometriotic lesions were induced by endometrial tissue transplantation into dorsal skinfold chambers of C57BL/6 mice, which were treated with the γ-secretase inhibitor DAPT or vehicle. Vascularization, morphology, and proliferation of the newly developing lesions were analyzed using intravital fluorescence microscopy, histology, and immunohistochemistry over 14 days. Inhibition of Notch signaling by DAPT significantly increased the number of angiogenic sprouts within the endometrial grafts during the first days after transplantation when compared to vehicle-treated controls. This was associated with an accelerated vascularization, as indicated by a higher functional microvessel density of DAPT-treated lesions on day 6. However, inhibition of Notch signaling did not affect the morphology and proliferating activity of the lesions, as previously described for tumors. Both DAPT- and vehicle-treated lesions finally consisted of cyst-like dilated glands, which were surrounded by a well-vascularized stroma and contained comparable numbers of proliferating cell nuclear antigen-positive cells. These findings demonstrate that sprouting angiogenesis in endometriotic lesions is controlled by Notch signaling. However, inhibition of Notch signaling does not have beneficial therapeutic effects on lesion development. Similar content being viewed by others

Galle PC (1989) Clinical presentation and diagnosis of endometriosis. Obstet Gynecol Clin North Am 16:29–42 Viganò P, Parazzini F, Somigliana E, Vercellini P (2004) Endometriosis: epidemiology and aetiological factors. Best Pract Res Clin Obstet Gynaecol 18:177–200 Sampson JA (1927) Peritoneal endometriosis due to menstrual dissemination of endometrial tissues into the peritoneal cavity. Am J Obstet Gynecol 14:422–469 Groothuis PG, Nap AW, Winterhager E, Grümmer R (2005) Vascular development in endometriosis. Angiogenesis 8:147–156 Laschke MW, Menger MD (2007) In vitro and in vivo approaches to study angiogenesis in the pathophysiology and therapy of endometriosis. Hum Reprod Update 13:331–342 Laschke MW, Schwender C, Scheuer C, Vollmar B, Menger MD (2008) Epigallocatechin-3-gallate inhibits estrogen-induced activation of endometrial cells in vitro and causes regression of endometriotic lesions in vivo. Hum Reprod 23:2308–2318 Taylor RN, Yu J, Torres PB, Schickedanz AC, Park JK, Mueller MD, Sidell N (2009) Mechanistic and therapeutic implications of angiogenesis in endometriosis. Reprod Sci 16:140–146 McLaren J (2000) Vascular endothelial growth factor and endometriotic angiogenesis. Hum Reprod Update 6:45–55 Taylor RN, Lebovic DI, Mueller MD (2002) Angiogenic factors in endometriosis. Ann N Y Acad Sci 955:89–100 Hellström M, Phng LK, Hofmann JJ, Wallgard E, Coultas L, Lindblom P, Alva J, Nilsson AK, Karlsson L, Gaiano N, Yoon K, Rossant J, Iruela-Arispe ML, Kalén M, Gerhardt H, Betsholtz C (2007) Dll4 signalling through Notch1 regulates formation of tip cells during angiogenesis. Nature 445:776–780 Zhang P, Yan X, Chen Y, Yang Z, Han H (2014) Notch signaling in blood vessels: from morphogenesis to homeostasis. Sci China Life Sci 57:774–780 Artavanis-Tsakonas S, Rand MD, Lake RJ (1999) Notch signaling: cell fate control and signal integration in development. Science 284:770–776 Sainson RC, Harris AL (2007) Anti-Dll4 therapy: can we block tumour growth by increasing angiogenesis? Trends Mol Med 13:389–395 Purow B (2012) Notch inhibition as a promising new approach to cancer therapy. Adv Exp Med Biol 727:305–319 Ridgway J, Zhang G, Wu Y, Stawicki S, Liang WC, Chanthery Y, Kowalski J, Watts RJ, Callahan C, Kasman I, Singh M, Chien M, Tan C, Hongo JA, de Sauvage F, Plowman G, Yan M (2006) Inhibition of Dll4 signalling inhibits tumour growth by deregulating angiogenesis. Nature 444:1083–1087 Noguera-Troise I, Daly C, Papadopoulos NJ, Coetzee S, Boland P, Gale NW, Lin HC, Yancopoulos GD, Thurston G (2006) Blockade of Dll4 inhibits tumour growth by promoting non-productive angiogenesis. Nature 444:1032–1037 Nenicu A, Körbel C, Gu Y, Menger MD, Laschke MW (2014) Combined blockade of angiotensin II type 1 receptor and activation of peroxisome proliferator-activated receptor-γ by telmisartan effectively inhibits vascularization and growth of murine endometriosis-like lesions. Hum Reprod 29:1011–1024 Laschke MW, Vorsterman van Oijen AE, Scheuer C, Menger MD (2011) In vitro and in vivo evaluation of the anti-angiogenic actions of 4-hydroxybenzyl alcohol. Br J Pharmacol 163:835–844 Laschke MW, Menger MD (2016) The dorsal skinfold chamber: a versatile tool for preclinical research in tissue engineering and regenerative medicine. Eur Cell Mater 32:202–215 Feng D, Menger MD, Wang H, Laschke MW (2014) Luminal epithelium in endometrial fragments affects their vascularization, growth and morphological development into endometriosis-like lesions in mice. Dis Model Mech 7:225–232 Sjölund J, Johansson M, Manna S, Norin C, Pietras A, Beckman S, Nilsson E, Ljungberg B, Axelson H (2008) Suppression of renal cell carcinoma growth by inhibition of Notch signaling in vitro and in vivo. J Clin Invest 118:217–228 Novella-Maestre E, Carda C, Noguera I, Ruiz-Saurí A, García-Velasco JA, Simón C, Pellicer A (2009) Dopamine agonist administration causes a reduction in endometrial implants through modulation of angiogenesis in experimentally induced endometriosis. Hum Reprod 24:1025–1035 Feng D, Welker S, Körbel C, Rudzitis-Auth J, Menger MD, Montenarh M, Laschke MW (2012) Protein kinase CK2 is a regulator of angiogenesis in endometriotic lesions. Angiogenesis 15:243–252 Feng D, Menger MD, Laschke MW (2013) Vascular disrupting effects of combretastatin A4 phosphate on murine endometriotic lesions. Fertil Steril 100:1459–1467 Shweiki D, Itin A, Soffer D, Keshet E (1992) Vascular endothelial growth factor induced by hypoxia may mediate hypoxia-initiated angiogenesis. Nature 359:843–845 Mailhos C, Modlich U, Lewis J, Harris A, Bicknell R, Ish-Horowicz D (2001) Delta4, an endothelial specific notch ligand expressed at sites of physiological and tumor angiogenesis. Differentiation 69:135–144 Gerhardt H, Golding M, Fruttiger M, Ruhrberg C, Lundkvist A, Abramsson A, Jeltsch M, Mitchell C, Alitalo K, Shima D, Betsholtz C (2003) VEGF guides angiogenic sprouting utilizing endothelial tip cell filopodia. J Cell Biol 161:1163–1177 Lobov IB, Renard RA, Papadopoulos N, Gale NW, Thurston G, Yancopoulos GD, Wiegand SJ (2007) Delta-like ligand 4 (Dll4) is induced by VEGF as a negative regulator of angiogenic sprouting. Proc Natl Acad Sci USA 104:3219–3224 Tung JJ, Tattersall IW, Kitajewski J (2012) Tips, stalks, tubes: notch-mediated cell fate determination and mechanisms of tubulogenesis during angiogenesis. Cold Spring Harb Perspect Med 2:a006601 Scehnet JS, Jiang W, Kumar SR, Krasnoperov V, Trindade A, Benedito R, Djokovic D, Borges C, Ley EJ, Duarte A, Gill PS (2007) Inhibition of Dll4-mediated signaling induces proliferation of immature vessels and results in poor tissue perfusion. Blood 109:4753–4760 Thurston G, Noguera-Troise I, Yancopoulos GD (2007) The delta paradox: DLL4 blockade leads to more tumour vessels but less tumour growth. Nat Rev Cancer 7:327–331 Becker CM, Beaudry P, Funakoshi T, Benny O, Zaslavsky A, Zurakowski D, Folkman J, D’Amato RJ, Ryeom S (2011) Circulating endothelial progenitor cells are up-regulated in a mouse model of endometriosis. Am J Pathol 178:1782–1791 Laschke MW, Giebels C, Menger MD (2011) Vasculogenesis: a new piece of the endometriosis puzzle. Hum Reprod Update 17:628–636 Laschke MW, Giebels C, Nickels RM, Scheuer C, Menger MD (2011) Endothelial progenitor cells contribute to the vascularization of endometriotic lesions. Am J Pathol 178:442–450 Rudzitis-Auth J, Nenicu A, Nickels RM, Menger MD, Laschke MW (2016) Estrogen stimulates homing of endothelial progenitor cells to endometriotic lesions. Am J Pathol 186:2129–2142 Laschke MW, Elitzsch A, Vollmar B, Vajkoczy P, Menger MD (2006) Combined inhibition of vascular endothelial growth factor (VEGF), fibroblast growth factor and platelet-derived growth factor, but not inhibition of VEGF alone, effectively suppresses angiogenesis and vessel maturation in endometriotic lesions. Hum Reprod 21:262–268

We are grateful for the excellent technical assistance of Janine Becker, Ruth M. Nickels, Julia Parakenings, and Sandra Schuler (Institute for Clinical and Experimental Surgery, Saarland University, Germany). Author information Authors and Affiliations Corresponding author Ethics declarations Conflict of interest The authors declare that they have no conflict of interest. Ethical approval All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures performed in studies involving animals were in accordance with the ethical standards of the institution or practice at which the studies were conducted. This article does not contain any studies with human participants performed by any of the authors. Rights and permissions About this article Cite this article Körbel, C., Gerstner, M.D., Menger, M.D. et al. Notch signaling controls sprouting angiogenesis of endometriotic lesions. Angiogenesis 21, 37–46 (2018). https://doi.org/10.1007/s10456-017-9580-7 Received: Accepted: Published: Issue date: DOI: https://doi.org/10.1007/s10456-017-9580-7