Skip to main content
SpringerLink
Log in

Extracellular matrix stiffness controls VEGF165 secretion and neuroblastoma angiogenesis via the YAP/RUNX2/SRSF1 axis

  • Original Paper
  • Published:
Angiogenesis Aims and scope Submit manuscript

Abstract

Aberrant variations in angiogenesis have been observed in tumor tissues with abnormal stiffness of extracellular matrix (ECM). However, it remains largely unclear how ECM stiffness influences tumor angiogenesis. Numerous studies have reported that vascular endothelial growth factor-A (VEGF-A) released from tumor cells plays crucial roles in angiogenesis. Hence, we demonstrated the role of ECM stiffness in VEGF-A release from neuroblastoma (NB) cells and the underlying mechanisms. Based on 17 NB clinical samples, a negative correlation was observed between the length of blood vessels and stiffness of NB tissues. In vitro, an ECM stiffness of 30 kPa repressed the secretion of VEGF165 from NB cells which subsequently inhibited the tube formation of human umbilical vein endothelial cells (HUVECs). Knocked down VEGF165 in NB cells or blocked VEGF165 with neutralizing antibodies both repressed the tube formation of HUVECs. Specifically, 30 kPa ECM stiffness repressed the expression and nuclear accumulation of Yes-associated protein (YAP) to regulate the expression of Serine/Arginine Splicing Factor 1 (SRSF1) via Runt-related transcription factor 2 (RUNX2), which may then subsequently induce the expression and secretion of VEGF165 in NB tumor cells. Through implantation of 3D col-Tgels with different stiffness into nude mice, the inhibitory effect of 30 kPa on NB angiogenesis was confirmed in vivo. Furthermore, we found that the inhibitory effect of 30 kPa stiffness on NB angiogenesis was reversed by YAP overexpression, suggesting the important role of YAP in NB angiogenesis regulated by ECM stiffness. Overall, our work not only showed a regulatory effect of ECM stiffness on NB angiogenesis, but also revealed a new signaling axis, YAP-RUNX2-SRSF1, that mediates angiogenesis by regulating the expression and secretion of VEGF165 from NB cells. ECM stiffness and the potential molecules revealed in the present study may be new therapeutic targets for NB angiogenesis.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4
Fig. 5
Fig. 6
Fig. 7

Similar content being viewed by others

References

  1. Ambros PF, Ambros IM, Brodeur GM et al (2009) International consensus for neuroblastoma molecular diagnostics: report from the International Neuroblastoma Risk Group (INRG) Biology Committee. Br J Cancer 100(9):1471–1482

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Cox TR, Erler JT (2011) Remodeling and homeostasis of the extracellular matrix: implications for fibrotic diseases and cancer. Dis Model Mech 4(2):165–178

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  3. Paszek MJ, Zahir N, Johnson KR et al (2005) Tensional homeostasis and the malignant phenotype. Cancer Cell 8(3):241–254

    Article  CAS  PubMed  Google Scholar 

  4. Engler AJ, Sen S, Sweeney HL et al (2006) Matrix elasticity directs stem cell lineage specification. Cell 126(4):677–689

    Article  CAS  PubMed  Google Scholar 

  5. Handorf AM, Zhou YX, Halanski MA et al (2015) Tissue stiffness dictates development, homeostasis, and disease progression. Organogenesis 11(1):1–15

    Article  PubMed  PubMed Central  Google Scholar 

  6. Meng ZP, Qiu YJ, Lin KC et al (2018) RAP2 mediates mechanoresponses of the Hippo pathway. Nature 560(7720):655–660

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Jeong J, Keum S, Kim D et al (2018) Spindle pole body component 25 homolog expressed by ECM stiffening is required for lung cancer cell proliferation. Biochem Biophys Res Commun 500(4):937–943

    Article  CAS  PubMed  Google Scholar 

  8. Pathak A, Kumar S (2012) Independent regulation of tumor cell migration by matrix stiffness and confinement. Proc Natl Acad Sci USA 109(26):10334–10339

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Peng YT, Chen ZY, Chen Y et al (2019) ROCK isoforms differentially modulate cancer cell motility by mechanosensing the substrate stiffness. Acta Biomater 88:86–101

    Article  CAS  PubMed  Google Scholar 

  10. Lam WA, Cao LZ, Umesh V et al (2010) Extracellular matrix rigidity modulates neuroblastoma cell differentiation and N-myc expression. Mol Cancer 9:35

    Article  PubMed  PubMed Central  Google Scholar 

  11. Bordeleaua F, Masona BN, Lollis EM et al (2016) Matrix stiffening promotes a tumor vasculature phenotype. Proc Natl Acad Sci USA 114(3):492–497

    Article  Google Scholar 

  12. López MR, Trinh AL, Sobrino A et al (2017) Recapitulating the human tumor microenvironment: colon tumor-derived extracellular matrix promotes angiogenesis and tumor cell growth. Biomaterials 116:118–129

    Article  Google Scholar 

  13. Shen Y, Wang XH, Lu JY et al (2020) Reduction of liver metastasis stiffness improves response to bevacizumab in metastatic colorectal cancer. Cancer Cell 37(6):800–817

    Article  CAS  PubMed  Google Scholar 

  14. Shoval H, Bluman AK, Karniely YB et al (2017) Tumor cells and their crosstalk with endothelial cells in 3D spheroids. Sci Rep 7(1):10428

    Article  PubMed  PubMed Central  Google Scholar 

  15. Sarkar S, Peng CC, Tung YC (2020) Comparison of VEGF-A secretion from tumor cells under cellular stresses in conventional monolayer culture and microfluidic threedimensional spheroid models. PLoS ONE 15(11):e0240833

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  16. Jakovljević G, Жulić S, Stepan J et al (2009) Vascular endothelial growth factor in children with neuroblastoma: a retrospective analysis. J Exp Clin Cancer Res 28(1):143

    Article  PubMed  PubMed Central  Google Scholar 

  17. Chanthery YH, Gustafson WH, Itsara M et al (2012) Paracrine signaling through MYCN enhances tumor-vascular interactions in neuroblastoma. Sci Transl Med 4(115):115ra3

    Article  PubMed  PubMed Central  Google Scholar 

  18. Pasquale MD, Castellano A, Sio L et al (2011) Bevacizumab in pediatric patients: how safe is it? Anticancer Res 31(11):3953–3957

    PubMed  Google Scholar 

  19. Geretti E, Shimizu A, Klagsbrun M (2008) Neuropilin structure governs VEGF and semaphorin binding and regulates angiogenesis. Angiogenesis 11(1):31–39

    Article  CAS  PubMed  Google Scholar 

  20. Mattei G, Gruca G, Rijnveld N et al (2015) The nano-epsilon dot method for strain rate viscoelastic characterisation of soft biomaterials by spherical nano-indentation. J Mech Behav Biomed Mater 50:150–159

    Article  CAS  PubMed  Google Scholar 

  21. Poursaleh A, Esfandiari G, Beigee FS et al (2019) Isolation of intimal endothelial cells from the human thoracic aorta: study protocol. Med J Islam Repub Iran 33:51

    PubMed  PubMed Central  Google Scholar 

  22. Tse JR, Engler AJ (2010) Preparation of Hydrogel substrates with tunable mechanical properties. Curr Protoc Cell Biol Chap 10: Unit. 10.16

  23. Artola AE, Andreu I, Beedle AEM et al (2017) Force tiggers YAP nuclear entry by regulating transport across nuclear pores. Cell 171(6):1397–1410

    Article  Google Scholar 

  24. Matsuzaki S, Canis M, Pouly JL et al (2015) Soft matrices inhibit cell proliferation and inactivate the fibrotic phenotype of deep endometriotic stromal cells in vitro. Hum Reprod 31(3):541–553

    Article  Google Scholar 

  25. James GC, Melissa G, Gopinath D et al (2015) The carboxyl terminus of VEGF-A is a potential target for anti-angiogenic therapy. Angiogenesis 18(1):23–30

    Article  Google Scholar 

  26. WagnerKD, Maï ME, Michael L et al (2019) Altered VEGF splicing isoform balance in tumor endothelium involves activation of splicing factors Srpk1 and Srsf1 by the Wilms’ tumor suppressor Wt1. Cells 8(1):41

    Article  Google Scholar 

  27. Dupont S, Morsut L, Aragona M et al (2011) Role of YAP/TAZ in mechanotransduction. Nature 474(7350):179–183

    Article  CAS  PubMed  Google Scholar 

  28. Dobrokhotov O, Samsonov M, Sokabe M et al (2018) Mechanoregulation and pathology of YAP/TAZ via Hippo and non-Hippo mechanisms. Clin Transl Med 7(1):23

    Article  PubMed  PubMed Central  Google Scholar 

  29. Kruik AM, Villasante A, Yaeger K et al (2018) Biomechanical regulation of drug sensitivity in an engineered model of human tumor. Biomaterials 150:150–161

    Article  Google Scholar 

  30. Han SY, Pang MF, Nelson CM et al (2018) Substratum stiffness tunes proliferation downstream of Wnt3a in part by regulating integrin-linked kinase and frizzled-1. J Cell Sci 131(8):jcs210476

    Article  PubMed  PubMed Central  Google Scholar 

  31. McKenzie AJ, Hicks SR, Svec KV et al (2018) The mechanical microenvironment regulates ovarian cancer cell morphology, migration, and spheroid disaggregation. Sci Rep 8(1):7228

    Article  PubMed  PubMed Central  Google Scholar 

  32. Tadeo I, Berbegall AP, Castel V et al (2016) Extracellular matrix composition defines an ultra-high-risk group of neuroblastoma within the high-risk patient cohort. Br J Cancer 115(4):480–489

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Abdeen AA, Weiss JB, Lee J et al (2014) Matrix composition and mechanics direct proangiogenic signaling from mesenchymal stem cells. Tissue Eng Part A 20(19–20):2737–2745

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Bandaru P, Cefaloni G, Vajhadin F et al (2020) Mechanical cues regulating proangiogenic potential of human mesenchymal stem cells through YAP-mediated. Mechanosens Small 16(25):e2001837

    Article  PubMed  Google Scholar 

  35. Mao L, Ding J, Zha YH (2011) HOXC9 links cell cycle exit and neuronal differentiation and is a prognostic marker in neuroblastoma. Cancer Res 71(12):4314–4324

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  36. Jogi A, Øraa I, Nilsson H et al (2003) Hypoxia-induced dedifferentiation in neuroblastoma cells. Cancer Lett 197(1–2):145–150

    Article  CAS  PubMed  Google Scholar 

  37. Vennin C, Chin VT, Warren SC et al (2017) Transient tissue priming via ROCK inhibition uncouples pancreatic cancer progression, sensitivity to chemotherapy, and metastasis. Sci Transl Med 9(384):eaai8504

    Article  PubMed  PubMed Central  Google Scholar 

  38. Joseph WF, Aaron BB, Vipul CC et al (2011) Stromal endothelial cells directly influence cancer progression. Sci Transl Med 3(66):66ra5

    Google Scholar 

  39. Duinen VV, Zhu D, Remakers C et al (2019) Perfused 3D angiogenic sprouting in a high-throughput in vitro platform. Angiogenesis 22(1):157–165

    Article  PubMed  Google Scholar 

  40. Michael RL, Steven JH, David OB (2007) Alternative splicing in angiogenesis: the vascular endothelial growth factor paradigm. Cancer Lett 249(2):133–142

    Article  Google Scholar 

  41. Arconde´ guy T, Lacazette E, Millevoi S (2013) VEGF-A mRNA processing, stability and translation: a paradigm for intricate regulation of gene expression at the post-transcriptional level. Nucleic Acids Res 41(17):7997–8010

    Article  Google Scholar 

  42. Maria PP (2012) The role of VEGF165b in pathophysiology. Cell Adhes Migr 6(6):1–8

    Google Scholar 

  43. Dawid GN, Elianna A, Emma SR et al (2010) Regulation of vascular endothelial growth factor (VEGF) splicing from pro-angiogenic to anti-angiogenic isoforms. J Biol Chem 285(8):5532–5540

    Article  Google Scholar 

  44. Shipra D, Adrian RK (2014) Emerging functions of SRSF1, splicing factor and oncoprotein, in RNA metabolism and cancer. Mol Cancer Res 12(9):1195–1204

    Article  Google Scholar 

  45. Zhou XX, Wang R, Li XB et al (2019) Splicing factor SRSF1 promotes gliomagenesis via oncogenic splice-switching of MYO1B. J Clin Invest 129(2):676–693

    Article  PubMed  PubMed Central  Google Scholar 

  46. Chen LL, Luo C, Shen L et al (2017) SRSF1 prevents DNA damage and promotes tumorigenesis through regulation of DBF4B PremRNA splicing. Cell Rep 21(12):3406–3413

    Article  CAS  PubMed  Google Scholar 

  47. Piccolo S, Dupont S, Cordenonsi M (2014) The biology of YAP/TAZ: hippo signaling and beyond. Physiol Rev 94(4):1287–1312

    Article  CAS  PubMed  Google Scholar 

  48. Iván MM, Georg H (2019) Hippo-YAP/TAZ signalling in organ regeneration and regenerative medicine. Nat Rev Mol Cell Biol 20(4):211–226

    Article  Google Scholar 

  49. John ML, Patrick S, Liu H et al (2012) The Hippo pathway target, YAP, promotes metastasis through its TEAD-interaction domain. Proc Natl Acad Sci USA 109(37):E2441–E2450

    Google Scholar 

  50. Jiang ZD, Zhou CC, Cheng L et al (2018) Inhibiting YAP expression suppresses pancreatic cancer progression by disrupting tumor-stromal interactions. J Exp Clin Cancer Res 37(1):69

    Article  PubMed  PubMed Central  Google Scholar 

  51. Liu ZJ, Yee PP, Wei YJ et al (2019) Differential YAP expression in glioma cells induces cell competition and promotes tumorigenesis. J Cell Sci 132(5):jcs225714

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  52. Lau AN, Curtis SJ, Fillmore CM et al (2014) Tumor-propagating cells and Yap/Taz activity contribute to lung tumor progression and metastasis. EMBO J 33(5):468–481

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  53. Lee TF, Tseng YC, Nguyen PA et al (2017) Enhanced YAP expression leads to EGFR TKI resistance in lung adenocarcinomas. Sci Rep 8(1):271

    Article  Google Scholar 

  54. Serrano I, McDonald PC, Lock F (2013) Inactivation of the Hippo tumour suppressor pathway by integrin-linked kinase. Nat Commun 4:2976

    Article  PubMed  Google Scholar 

  55. Pankova D, Jiang YY, Chatzifrangkeskou M et al (2019) RASSF1A controls tissue stiffness and cancer stem-like cells in lung adenocarcinoma. EMBO J 38(13):e100532

    Article  PubMed  PubMed Central  Google Scholar 

  56. Totaro A, Castellan M, Battilana G et al (2017) YAP/TAZ link cell mechanics to Notch signalling to control epidermal stem cell fate. Nat Commun 8:15206

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  57. Qin X, Lv XY, Li P et al (2020) Matrix stiffness modulates ILK-mediated YAP activation to control the drug resistance of breast cancer cells. Biochim Biophys Acta Mol Basis Dis 1866(3):165625

    Article  CAS  PubMed  Google Scholar 

  58. Barreto S, Vazquez AG, Cameron AR et al (2017) Identification of the mechanisms by which age alters the mechanosensitivity of mesenchymal stromal cells on substrates of differing stiffness: implications for osteogenesis and angiogenesis. Acta Biomater 53:59–69

    Article  PubMed  Google Scholar 

Download references

Acknowledgements

This research was supported by grants from the National Natural Science Foundation of China, No. 11625209.

Author information

Authors and Affiliations

  1. Institute of Mechanobiology & Medical Engineering, School of Life Sciences & Biotechnology, Shanghai Jiao Tong University, 800 Dongchuan Road, Minhang, 200240, Shanghai, China

    Min Bao, Yi Chen, Ji-Ting Liu, Han Bao, Wen-Bin Wang & Ying-Xin Qi

  2. Department of Pediatric Surgery, Xinhua Hospital, Shanghai Jiao Tong University School of Medicine, 1665 Kongjiang Road, Yangpu, Shanghai, 200092, China

    Fan Lv

Authors
  1. Min Bao
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Yi Chen
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Ji-Ting Liu
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Han Bao
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Wen-Bin Wang
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Ying-Xin Qi
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Fan Lv
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding authors

Correspondence to Ying-Xin Qi or Fan Lv.

Ethics declarations

Conflict of interest

The authors declare that they have no conflict of interest.

Additional information

Publisher’s Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.

Supplementary file 1 (DOCX 33865 KB)

Rights and permissions

Reprints and permissions

About this article

Check for updates. Verify currency and authenticity via CrossMark

Cite this article

Bao, M., Chen, Y., Liu, JT. et al. Extracellular matrix stiffness controls VEGF165 secretion and neuroblastoma angiogenesis via the YAP/RUNX2/SRSF1 axis. Angiogenesis 25, 71–86 (2022). https://doi.org/10.1007/s10456-021-09804-7

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10456-021-09804-7

Keywords

Search

Navigation