Elsevier

Urology

Volume 81, Issue 2, February 2013, Pages 424-431
Urology

Reconstructive Urology
Engineering of Corpus Cavernosum Using Vascular Endothelial Growth Factor-expressing Muscle-derived Stem Cells Seeded on Acellular Corporal Collagen Matrices

https://doi.org/10.1016/j.urology.2012.10.042Get rights and content

Objective

To explore the feasibility of developing vascularized tissue-engineered corpus cavernosum with vascular endothelial growth factor (VEGF)-expressing muscle-derived stem cells (MDSCs) as seed cells in a rabbit model.

Materials and Methods

MDSCs were isolated and expanded in vitro and transfected with human VEGF165 lentiviral gene vector. The release of VEGF was confirmed using enzyme-linked immunosorbent assay. Acellular corporal collagen matrices (ACCMs) were obtained from donor rabbit penile tissue using an established decellularization process. Transfected and untransfected MDSCs were seeded on ACCMs. Histochemistry and scanning electron microscopy were performed to analyze the growth and distribution of MDSCs. After constructing animal models, we transplanted the seeded ACCMs to the excised corporal space. The neocorpora was harvested and assayed with Western blot and immunohistochemistry at the first and second month.

Results

Enzyme-linked immunosorbent assay indicated that the release of VEGF was significantly increased in the MDSC-VEGF group compared with the other groups. Histochemistry and scanning electron microscopy indicated that VEGF-expressing MDSCs showed better growth and adequate attachment than other groups on the ACCMs in vitro. Immunohistochemical staining showed that the expression of α-smooth muscle actin, von Willebrand factor, and CD31 in the MDSC-VEGF group was markedly increased at different points. The MDSC-VEGF group showed greater improvement on cavernosal contractile function than the other groups. The expression of endothelial nitric oxide synthase in the engineered corporal tissues was significantly greater in the MDSC-VEGF group than in the other groups at different measurement points.

Conclusion

As seed cells, VEGF-overexpressing MDSCs could greatly increase the density of capillaries and the content of endothelial smooth muscle cells in constructing the engineered corporal tissues than can untransfected MDSCs.

Section snippets

Isolation and Expansion of MDSCs

The hind limb gastrocnemius muscles were removed from 2-week-old normal New Zealand white rabbits. MDSCs were obtained and purified by established preplate techniques.13, 19 The proliferation medium was Dulbecco’s modified Eagle’s medium (Gibco, Life Technologies, Carlsband, CA) with 10% fetal bovine serum (Gibco), 10% horse serum (Gibco), 0.5% chick embryo extract (Gibco), and 1% penicillin/streptomycin (Gibco).

Cell Transfection, MDSC Characterization, and VEGF Expression

The human VEGF165 lentiviral gene vector (LV-GFP-VEGF), which contained a green

Transfection of VEGF Gene into MDSCs

After lentiviral transfection, the expression of desmin, Bcl-2, CD34, and CD45, detected using flow cytometry, did not show any significant changes (Fig. 1A). Using fluorescence microscopy, GFP expression in the transfected MDSCs was observed, and transfection efficiency was detected by flow cytometry (Fig. 1B). The exper-iments with MDSCs were performed at a multiplicity of infection of 15. Western blot analysis was used to detected the protein expression of α-SMA in the MDSC, MDSC-vector, and

Comment

Cavernous smooth muscle cells and endothelial cells are the main components of the corpus cavernosum and play an important role in maintaining and regulating the function of the penis. Reconstitution of the corpus cavernosum smooth muscle and sinus endothelium are always the main research components of a TECC. To achieve fully functional tissue-engineered corporal tissue, various approaches have been studied, including a combination of seed cells with biodegradable polymer scaffolds10, 20 or

Conclusion

In the present study, we successfully constructed lentiviral-mediated VEGF-expressing MDSCs and demonstrated their role, as seed cells, in increasing the content of endothelial cells, smooth muscle cells, and capillaries in constructing the TECC. In particular, VEGF-expressing MDSCs greatly increased the capillary density in the engineered corporal tissue compared with simple MDSC. The combination of MDSC seeding with VEGF gene therapy can be considered as a potential new seed cell model to

References (28)

  • J.H. Hwang et al.

    Isolation of muscle derived stem cells from rat and its smooth muscle differentiation

    Mol Cells

    (2004)
  • G. Selvaqqi et al.

    Penile reconstruction/formation

    Curr Opin Urol

    (2008)
  • M. Doornaert et al.

    Penile reconstruction with the radial forearm flap: an update

    Handchir Mikrochir Plast Chir

    (2011)
  • M. Mutaf et al.

    A true one-stage nonmicrosurgical technique for total phallic reconstruction

    Ann Plast Surg

    (2006)
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    Financial Disclosure: The authors declare that they have no relevant financial interests.

    Funding Support: This work was supported by the National Natural Science Foundation of China (grant 30672183), the Science and Technology Plan of Guangdong Province (grant 1111220600060), and the Natural Science Foundation of Guangdong Province (grant 8151008901000202).

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