Molecular Biotechnology

, Volume 60, Issue 6, pp 396–411 | Cite as

The Methods and Mechanisms to Differentiate Endothelial-Like Cells and Smooth Muscle Cells from Mesenchymal Stem Cells for Vascularization in Vaginal Reconstruction

  • Hua Zhang
  • Jingkun Zhang
  • Xianghua Huang
  • Yanan Li
Original Paper


Endothelial cells and smooth muscle cells (SMCs) are important aspects of vascularization in vaginal reconstruction. Research has confirmed that mesenchymal stem cells could differentiate into endothelial-like cells and SMCs. But the methods were more complicated and the mechanism was unknown. In the current study, we induced the bone mesenchymal stem cells (BMSCs) to differentiate into endothelial-like cells and SMCs in vitro by differentiation medium and investigated the effect of Wnt/β-catenin signaling on the differentiation process of BMSCs. Results showed that the hypoxic environment combined with VEGF and bFGF could induce increased expression of endothelial-like cells markers VEGFR1, VEGFR2, and vWF. The SMCs derived from BMSCs induced by TGF-β1 and PDGF-AB significantly expressed SMC markers SMMHC11 and α-SMA. The data also showed that activation of Wnt/β-catenin signaling could promote the differentiation of BMSCs into endothelial-like cells and SMCs. Thus, we established endothelial-like cells and SMCs in vitro by more simple methods, presented the important role of hypoxic environment on the differentiation of BMSCs into endothelial-like cells, and confirmed that the Wnt/β-catenin signaling pathway has a positive impact on the differentiation of BMSCs into endothelial-like cells and SMCs. This is important for vascular reconstruction.


Cell differentiation Endothelial-like cells Hypoxic environment Mesenchymal stem cell Smooth muscle cells 



We thank Dr. Zhang and Dr. Xu of the University of Hebei Pharmacology Laboratory of Shijiazhuang, China, for their help with our research. This work was funded by the National Natural Science Foundation of China (Grant No. 81671407).

Compliance with Ethical Standards

Conflict of interest

The authors declare that they have no conflict of interest.


  1. 1.
    Varner, R. E., Younger, J. B., & Blackwell, R. E. (1985). Mullerian dysgenesis. The Journal of Reproductive Medicine, 30, 443–450.Google Scholar
  2. 2.
    Nakhal, R. S., & Creighton, S. M. (2012). Management of vaginal agenesis. Journal of Pediatric and Adolescent Gynecology, 25, 352–357.CrossRefGoogle Scholar
  3. 3.
    Idrees, M. T., Deligdisch, L., & Altchek, A. (2009). Squamous papilloma with hyperpigmentation in the skin graft of the neovagina in Rokitansky syndrome: Literature review of benign and malignant lesions of the neovagina. Journal of Pediatric and Adolescent Gynecology, 22, e148–e155.CrossRefGoogle Scholar
  4. 4.
    Patra, S., & Young, V. (2016). A Review of 3D Printing Techniques and the Future in Biofabrication of Bioprinted Tissue. Cell Biochemistry and Biophysics, 74, 93.CrossRefGoogle Scholar
  5. 5.
    Zhang, J. K., Du, R. X., Zhang, L., Li, Y. N., Zhang, M. L., Zhao, S., et al. (2017). A new material for tissue engineered vagina reconstruction: Acellular porcine vagina matrix. Journal of Biomedical Materials Research, Part A, 105, 1949–1959.CrossRefGoogle Scholar
  6. 6.
    Poole, T. J., Finkelstein, E. B., & Cox, C. M. (2001). The role of FGF and VEGF in angioblast induction and migration during vascular development. Developmental Dynamics, 220, 1–17.CrossRefGoogle Scholar
  7. 7.
    Wang, C., Cen, L., Yin, S., Liu, Q., Liu, W., Cao, Y., et al. (2010). A small diameter elastic blood vessel wall prepared under pulsatile conditions from polyglycolic acid mesh and smooth muscle cells differentiated from adipose-derived stem cells. Biomaterials, 31, 621–630.CrossRefGoogle Scholar
  8. 8.
    Coll-Bonfill, N., Musri, M. M., Ivo, V., Barbera, J. A., & Tura-Ceide, O. (2015). Transdifferentiation of endothelial cells to smooth muscle cells play an important role in vascular remodelling. American Journal of Stem Cells, 4, 13–21.Google Scholar
  9. 9.
    Bara, J. J., Richards, R. G., Alini, M., & Stoddart, M. J. (2014). Concise review: Bone marrow-derived mesenchymal stem cells change phenotype following in vitro culture—Implications for basic research and the clinic. Stem Cells, 32, 1713–1723.CrossRefGoogle Scholar
  10. 10.
    Wei, X., Yang, X., Han, Z. P., Qu, F. F., Shao, L., & Shi, Y. F. (2013). Mesenchymal stem cells: A new trend for cell therapy. Acta Pharmacologica Sinica, 34, 747–754.CrossRefGoogle Scholar
  11. 11.
    De Becker, A., & Van Riet, I. (2015). Mesenchymal stromal cell therapy in hematology: From laboratory to clinic and back again. Stem Cells and Development, 24, 1713–1729.CrossRefGoogle Scholar
  12. 12.
    Fibbe, W. E., Nauta, A. J., & Roelofs, H. (2007). Modulation of immune responses by mesenchymal stem cells. Annals of the New York Academy of Sciences, 1106, 272–278.CrossRefGoogle Scholar
  13. 13.
    Dai, R., Wang, Z., Samanipour, R., Koo, K. I., & Kim, K. (2016). Adipose-derived stem cells for tissue engineering and regenerative medicine applications. Stem Cells International, 2016, 6737345.Google Scholar
  14. 14.
    Trohatou, O., Anagnou, N. P., & Roubelakis, M. G. (2013). Human amniotic fluid stem cells as an attractive tool for clinical applications. Current Stem Cell Research and Therapy, 8, 125–132.CrossRefGoogle Scholar
  15. 15.
    Flynn, A., Barry, F., & O’Brien, T. (2007). UC blood-derived mesenchymal stromal cells: An overview. Cytotherapy, 9, 717–726.CrossRefGoogle Scholar
  16. 16.
    Liu, R. M., Sun, R. G., Zhang, L. T., Zhang, Q. F., Chen, D. X., Zhong, J. J., et al. (2016). Hyaluronic acid enhances proliferation of human amniotic mesenchymal stem cells through activation of Wnt/beta-catenin signaling pathway. Experimental Cell Research, 345, 218–229.CrossRefGoogle Scholar
  17. 17.
    Mansson-Broberg, A., Rodin, S., Bulatovic, I., Ibarra, C., Lofling, M., Genead, R., et al. (2016). Wnt/beta-catenin stimulation and laminins support cardiovascular cell progenitor expansion from human fetal cardiac mesenchymal stromal cells. Stem Cell Reports, 6, 607–617.CrossRefGoogle Scholar
  18. 18.
    Li, Y., Liu, F., Zhang, Z., Zhang, M., Cao, S., Li, Y., et al. (2015). Bone marrow mesenchymal stem cells could acquire the phenotypes of epithelial cells and accelerate vaginal reconstruction combined with small intestinal submucosa. Cell Biology International, 39, 1225–1233.CrossRefGoogle Scholar
  19. 19.
    Oswald, J., Boxberger, S., Jorgensen, B., Feldmann, S., Ehninger, G., Bornhauser, M., et al. (2004). Mesenchymal stem cells can be differentiated into endothelial cells in vitro. Stem Cells, 22, 377–384.CrossRefGoogle Scholar
  20. 20.
    Ikhapoh, I. A., Pelham, C. J., & Agrawal, D. K. (2015). Sry-type HMG box 18 contributes to the differentiation of bone marrow-derived mesenchymal stem cells to endothelial cells. Differentiation: Research in Biological Diversity, 89, 87–96.CrossRefGoogle Scholar
  21. 21.
    Hasanzadeh, E., Amoabediny, G., Haghighipour, N., Gholami, N., Mohammadnejad, J., Shojaei, S., et al. (2017). The stability evaluation of mesenchymal stem cells differentiation toward endothelial cells by chemical and mechanical stimulation. In vitro cellular and developmental biology. Animal, 53, 818.Google Scholar
  22. 22.
    Zhu, Z., Gan, X., Fan, H., & Yu, H. (2015). Mechanical stretch endows mesenchymal stem cells stronger angiogenic and anti-apoptotic capacities via NFkappaB activation. Biochemical and Biophysical Research Communications, 468, 601–605.CrossRefGoogle Scholar
  23. 23.
    Zhang, R., Wang, N., Zhang, L. N., Huang, N., Song, T. F., Li, Z. Z., et al. (2016). Knockdown of DNMT1 and DNMT3a promotes the angiogenesis of human mesenchymal stem cells leading to arterial specific differentiation. Stem Cells, 34, 1273–1283.CrossRefGoogle Scholar
  24. 24.
    Lee, J. H., Ryu, J. M., Han, Y. S., Zia, M. F., Kwon, H. Y., Noh, H., et al. (2016). Fucoidan improves bioactivity and vasculogenic potential of mesenchymal stem cells in murine hind limb ischemia associated with chronic kidney disease. Journal of Molecular and Cellular Cardiology, 97, 169–179.CrossRefGoogle Scholar
  25. 25.
    Efimenko, A., Sagaradze, G., Akopyan, Z., Lopatina, T., & Kalinina, N. (2016). Data supporting that miR-92a suppresses angiogenic activity of adipose-derived mesenchymal stromal cells by down-regulating hepatocyte growth factor. Data in Brief, 6, 295–310.CrossRefGoogle Scholar
  26. 26.
    Aji, K., Maimaijiang, M., Aimaiti, A., Rexiati, M., Azhati, B., Tusong, H., et al. (2016). Differentiation of human adipose derived stem cells into smooth muscle cells is modulated by CaMKIIgamma. Stem Cells International, 2016, 1267480.CrossRefGoogle Scholar
  27. 27.
    Alimperti, S., You, H., George, T., Agarwal, S. K., & Andreadis, S. T. (2014). Cadherin-11 regulates both mesenchymal stem cell differentiation into smooth muscle cells and the development of contractile function in vivo. Journal of Cell Science, 127, 2627–2638.CrossRefGoogle Scholar
  28. 28.
    Brun, J., Abruzzese, T., Rolauffs, B., Aicher, W. K., & Hart, M. L. (2016). Choice of xenogenic-free expansion media significantly influences the myogenic differentiation potential of human bone marrow-derived mesenchymal stromal cells. Cytotherapy, 18, 344–359.CrossRefGoogle Scholar
  29. 29.
    Jeon, E. S., Moon, H. J., Lee, M. J., Song, H. Y., Kim, Y. M., Bae, Y. C., et al. (2006). Sphingosylphosphorylcholine induces differentiation of human mesenchymal stem cells into smooth-muscle-like cells through a TGF-beta-dependent mechanism. Journal of Cell Science, 119, 4994–5005.CrossRefGoogle Scholar
  30. 30.
    Lin, L., Qiu, Q., Zhou, N., Dong, W., Shen, J., Jiang, W., et al. (2016). Dickkopf-1 is involved in BMP9-induced osteoblast differentiation of C3H10T1/2 mesenchymal stem cells. BMB Reports, 49, 179–184.CrossRefGoogle Scholar
  31. 31.
    Chu, E. Y., Hens, J., Andl, T., Kairo, A., Yamaguchi, T. P., Brisken, C., et al. (2004). Canonical WNT signaling promotes mammary placode development and is essential for initiation of mammary gland morphogenesis. Development, 131, 4819–4829.CrossRefGoogle Scholar
  32. 32.
    Lee, D. E., Ayoub, N., & Agrawal, D. K. (2016). Mesenchymal stem cells and cutaneous wound healing: Novel methods to increase cell delivery and therapeutic efficacy. Stem Cell Research and Therapy, 7, 37.CrossRefGoogle Scholar
  33. 33.
    Khan, S., Villalobos, M. A., Choron, R. L., Chang, S., Brown, S. A., Carpenter, J. P., et al. (2016). Fibroblast growth factor and vascular endothelial growth factor play a critical role in endotheliogenesis from human adipose-derived stem cells. Journal of Vascular Surgery, 65, 1483.CrossRefGoogle Scholar
  34. 34.
    Brun, J., Lutz, K. A., Neumayer, K. M., Klein, G., Seeger, T., Uynuk-Ool, T., et al. (2015). Smooth muscle-like cells generated from human mesenchymal stromal cells display marker gene expression and electrophysiological competence comparable to bladder smooth muscle cells. PLoS ONE, 10, e0145153.CrossRefGoogle Scholar
  35. 35.
    Allameh, A., Jazayeri, M., & Adelipour, M. (2016). In vivo vascularization of endothelial cells derived from bone marrow mesenchymal stem cells in SCID mouse model. Cell Journal, 18, 179–188.Google Scholar
  36. 36.
    Amorim, B. R., Silverio, K. G., Casati, M. Z., Sallum, E. A., Kantovitz, K. R., & Nociti, F. H., Jr. (2016). Neuropilin controls endothelial differentiation by mesenchymal stem cells from the periodontal ligament. Journal of Periodontology, 87, e138–e147.CrossRefGoogle Scholar
  37. 37.
    Fathi, F., Rezabakhsh, A., Rahbarghazi, R., & Rashidi, M. R. (2017). Early-stage detection of VE-cadherin during endothelial differentiation of human mesenchymal stem cells using SPR biosensor. Biosensors and Bioelectronics, 96, 358–366.CrossRefGoogle Scholar
  38. 38.
    Shin, J. W., Park, S. H., Kang, Y. G., Wu, Y., Choi, H. J., & Shin, J. W. (2016). Changes, and the relevance thereof, in mitochondrial morphology during differentiation into endothelial cells. PLoS ONE, 11, e0161015.CrossRefGoogle Scholar
  39. 39.
    Robins, J. C., Akeno, N., Mukherjee, A., Dalal, R. R., Aronow, B. J., Koopman, P., et al. (2005). Hypoxia induces chondrocyte-specific gene expression in mesenchymal cells in association with transcriptional activation of Sox9. Bone, 37, 313–322.CrossRefGoogle Scholar
  40. 40.
    Hung, S. P., Ho, J. H., Shih, Y. R., Lo, T., & Lee, O. K. (2012). Hypoxia promotes proliferation and osteogenic differentiation potentials of human mesenchymal stem cells. Journal of Orthopaedic Research, 30, 260–266.CrossRefGoogle Scholar
  41. 41.
    Giaccia, A. J., Simon, M. C., & Johnson, R. (2004). The biology of hypoxia: The role of oxygen sensing in development, normal function, and disease. Genes and Development, 18, 2183–2194.CrossRefGoogle Scholar
  42. 42.
    Roemeling-van Rhijn, M., Mensah, F. K., Korevaar, S. S., Leijs, M. J., van Osch, G. J., Ijzermans, J. N., et al. (2013). Effects of hypoxia on the immunomodulatory properties of adipose tissue-derived mesenchymal stem cells. Frontiers in Immunology, 4, 203.CrossRefGoogle Scholar
  43. 43.
    Tsai, C. C., Chen, Y. J., Yew, T. L., Chen, L. L., Wang, J. Y., Chiu, C. H., et al. (2011). Hypoxia inhibits senescence and maintains mesenchymal stem cell properties through down-regulation of E2A-p21 by HIF-TWIST. Blood, 117, 459–469.CrossRefGoogle Scholar
  44. 44.
    Chow, D. C., Wenning, L. A., Miller, W. M., & Papoutsakis, E. T. (2001). Modeling pO(2) distributions in the bone marrow hematopoietic compartment. II. Modified Kroghian models. Biophysical Journal, 81, 685–696.CrossRefGoogle Scholar
  45. 45.
    Talks, K. L., Turley, H., Gatter, K. C., Maxwell, P. H., Pugh, C. W., Ratcliffe, P. J., et al. (2000). The expression and distribution of the hypoxia-inducible factors HIF-1alpha and HIF-2alpha in normal human tissues, cancers, and tumor-associated macrophages. The American Journal of Pathology, 157, 411–421.CrossRefGoogle Scholar
  46. 46.
    Duval, E., Bauge, C., Andriamanalijaona, R., Benateau, H., Leclercq, S., Dutoit, S., et al. (2012). Molecular mechanism of hypoxia-induced chondrogenesis and its application in in vivo cartilage tissue engineering. Biomaterials, 33, 6042–6051.CrossRefGoogle Scholar
  47. 47.
    Cicione, C., Muinos-Lopez, E., Hermida-Gomez, T., Fuentes-Boquete, I., Diaz-Prado, S., & Blanco, F. J. (2013). Effects of severe hypoxia on bone marrow mesenchymal stem cells differentiation potential. Stem Cells International, 2013, 232896.CrossRefGoogle Scholar
  48. 48.
    Holzwarth, C., Vaegler, M., Gieseke, F., Pfister, S. M., Handgretinger, R., Kerst, G., et al. (2010). Low physiologic oxygen tensions reduce proliferation and differentiation of human multipotent mesenchymal stromal cells. BMC Cell Biology, 11, 11.CrossRefGoogle Scholar
  49. 49.
    Pacary, E., Legros, H., Valable, S., Duchatelle, P., Lecocq, M., Petit, E., et al. (2006). Synergistic effects of CoCl(2) and ROCK inhibition on mesenchymal stem cell differentiation into neuron-like cells. Journal of Cell Science, 119, 2667–2678.CrossRefGoogle Scholar
  50. 50.
    Wang, X., Liu, C., Li, S., Xu, Y., Chen, P., Liu, Y., et al. (2015). Hypoxia precondition promotes adipose-derived mesenchymal stem cells based repair of diabetic erectile dysfunction via augmenting angiogenesis and neuroprotection. PLoS ONE, 10, e0118951.CrossRefGoogle Scholar
  51. 51.
    Xing, Y., Hou, J., Guo, T., Zheng, S., Zhou, C., Huang, H., et al. (2014). microRNA-378 promotes mesenchymal stem cell survival and vascularization under hypoxic-ischemic conditions in vitro. Stem Cell Research and Therapy, 5, 130.CrossRefGoogle Scholar
  52. 52.
    Tian, H., Bharadwaj, S., Liu, Y., Ma, H., Ma, P. X., Atala, A., et al. (2010). Myogenic differentiation of human bone marrow mesenchymal stem cells on a 3D nano fibrous scaffold for bladder tissue engineering. Biomaterials, 31, 870–877.CrossRefGoogle Scholar
  53. 53.
    Narita, Y., Yamawaki, A., Kagami, H., Ueda, M., & Ueda, Y. (2008). Effects of transforming growth factor-beta 1 and ascorbic acid on differentiation of human bone-marrow-derived mesenchymal stem cells into smooth muscle cell lineage. Cell and Tissue Research, 333, 449–459.CrossRefGoogle Scholar
  54. 54.
    Beamish, J. A., He, P., Kottke-Marchant, K., & Marchant, R. E. (2010). Molecular regulation of contractile smooth muscle cell phenotype: Implications for vascular tissue engineering. Tissue Engineering. Part B, Reviews, 16, 467–491.CrossRefGoogle Scholar
  55. 55.
    Cadigan, K. M., & Nusse, R. (1997). Wnt signaling: A common theme in animal development. Genes and Development, 11, 3286–3305.CrossRefGoogle Scholar
  56. 56.
    Wodarz, A., & Nusse, R. (1998). Mechanisms of Wnt signaling in development. Annual Review of Cell and Developmental Biology, 14, 59–88.CrossRefGoogle Scholar
  57. 57.
    Novak, A., & Dedhar, S. (1999). Signaling through beta-catenin and Lef/Tcf. Cellular and Molecular Life Sciences: CMLS, 56, 523–537.CrossRefGoogle Scholar
  58. 58.
    Sun, J., Chen, J., Cao, J., Li, T., Zhuang, S., & Jiang, X. (2016). IL-1beta-stimulated beta-catenin up-regulation promotes angiogenesis in human lung-derived mesenchymal stromal cells through a NF-kappaB-dependent microRNA-433 induction. Oncotarget, 7, 59429–59440.Google Scholar
  59. 59.
    Sun, Z., Wang, C., Shi, C., Sun, F., Xu, X., Qian, W., et al. (2014). Activated Wnt signaling induces myofibroblast differentiation of mesenchymal stem cells, contributing to pulmonary fibrosis. International Journal of Molecular Medicine, 33, 1097–1109.CrossRefGoogle Scholar
  60. 60.
    van der Horst, G., van der Werf, S. M., Farih-Sips, H., van Bezooijen, R. L., Lowik, C. W., & Karperien, M. (2005). Downregulation of Wnt signaling by increased expression of Dickkopf-1 and -2 is a prerequisite for late-stage osteoblast differentiation of KS483 cells. Journal of Bone and Mineral Research, 20, 1867–1877.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC, part of Springer Nature 2018

Authors and Affiliations

  • Hua Zhang
    • 1
  • Jingkun Zhang
    • 1
  • Xianghua Huang
    • 1
  • Yanan Li
    • 1
  1. 1.Department of Obstetrics and GynecologyThe Second Hospital of Hebei Medical UniversityShijiazhuangChina

Personalised recommendations