Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Original Article
  • Published:

A coculture system of cavernous endothelial and smooth muscle cells

International Journal of Impotence Research volume 25, pages 63–68 (2013)Cite this article

Subjects

Abstract

In erectile dysfunction (ED) research, monocultures of cavernous endothelial cells (CECs) and smooth muscle cells (CSMCs) have been reported, but a CEC–CSMC coculture system is still lacking. In the present study, we wished to investigate the feasibility of setting up such a system and test whether it can be used for diabetic ED research. Cavernous tissues were obtained from patients undergoing surgery for penile prosthesis. CSMCs were isolated by explant culture and verified by calponin staining. CECs were isolated by binding to CD31 antibody, followed by magnetic capture. These CECs were nearly 100% pure endothelial cells as determined by flow cytometric analysis for endothelial markers CD31, vWF and eNOS. Functional analyses, that is, low-density lipoprotein (LDL) uptake and capillary tube formation, also confirmed their endothelial phenotype. When cocultured with CSMCs, CECs formed capillary-like structures, and based on the extent of this capillary-like network, it was determined that a ratio of 1:4 in cell number between CECs and CSMCs was better than ratios of 1:1 and 1:9. It was also found that direct contact between CECs and CSMCs was necessary and a coculture period of 3 weeks was optimal. Autologous CSMCs were better than allogeneic CSMCs, and fibroblasts were completely incompetent. When treated with high glucose (25 mM), the CEC–CSMC coculture expressed significantly lower level of CD31 but significantly higher level of collagen-IV (Col-IV), and the diameter of the capillaries increased significantly, when compared with normal glucose (5 mM)-treated cocultures. These data are consistent with previously observed changes in the cavernous tissues of diabetic patients and thus suggest that the coculture system could be utilized for diabetic ED research.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Figure 1
Figure 2
Figure 3
Figure 4
Figure 5

Similar content being viewed by others

References

  1. Lin G, Qiu X, Fandel TM, Albersen M, Wang Z, Lue TF et al. Improved penile histology by phalloidin stain: circular and longitudinal cavernous smooth muscles, dual-endothelium arteries, and erectile dysfunction-associated changes. Urology 2011; 78: 970.e1–8.

    Article  Google Scholar 

  2. Gondre M, Christ GJ . Endothelin-1-induced alterations in phenylephrine-induced contractile responses are largely additive in physiologically diverse rabbit vasculature. J Pharmacol Exp Ther 1998; 286: 635–642.

    CAS  PubMed  Google Scholar 

  3. Italiano G, Calabro A, Spini S, Ragazzi E, Pagano F . Functional response of cavernosal tissue to distension. Urol Res 1998; 26: 39–44.

    Article  CAS  PubMed  Google Scholar 

  4. Hurt KJ, Musicki B, Palese MA, Crone JK, Becker RE, Moriarity JL et al. Akt-dependent phosphorylation of endothelial nitric-oxide synthase mediates penile erection. Proc Natl Acad Sci USA 2002; 99: 4061–4066.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  5. Schuster A, Oishi H, Beny JL, Stergiopulos N, Meister JJ . Simultaneous arterial calcium dynamics and diameter measurements: application to myoendothelial communication. Am J Physiol Heart Circ Physiol 2001; 280: H1088–H1096.

    Article  CAS  PubMed  Google Scholar 

  6. Moncada S, Palmer RM, Higgs EA . Nitric oxide: physiology, pathophysiology, and pharmacology. Pharmacol Rev 1991; 43: 109–142.

    CAS  PubMed  Google Scholar 

  7. Niwa K, Kado T, Sakai J, Karino T . The effects of a shear flow on the uptake of LDL and acetylated LDL by an EC monoculture and an EC-SMC coculture. Ann Biomed Eng 2004; 32: 537–543.

    Article  PubMed  Google Scholar 

  8. Truskey GA . Endothelial cell vascular smooth muscle cell co-culture assay for high throughput screening assays for discovery of anti-angiogenesis agents and other therapeutic molecules. Int J High Throughput Screen 2010; 2010: 171–181.

    Article  PubMed  PubMed Central  Google Scholar 

  9. Powell RJ, Cronenwett JL, Fillinger MF, Wagner RJ, Sampson LN . Endothelial cell modulation of smooth muscle cell morphology and organizational growth pattern. Ann Vasc Surg 1996; 10: 4–10.

    Article  CAS  PubMed  Google Scholar 

  10. Lin CS, Lin G, Wang Z, Maddah SA, Lue TF . Upregulation of monocyte chemoattractant protein 1 and effects of transforming growth factor-beta 1 in Peyronie’s disease. Biochem Biophys Res Commun 2002; 295: 1014–1019.

    Article  CAS  PubMed  Google Scholar 

  11. Ning H, Lin G, Lue TF, Lin CS . Effects of estrogen, raloxifene, and levormeloxifene on the expression of Rho-kinase signaling molecules in urethral smooth muscle cells. Urology 2011; 76: 1517.e6–11.

    Google Scholar 

  12. van Beijnum JR, Rousch M, Castermans K, van der Linden E, Griffioen AW . Isolation of endothelial cells from fresh tissues. Nat Protoc 2008; 3: 1085–1091.

    Article  CAS  PubMed  Google Scholar 

  13. Evensen L, Micklem DR, Blois A, Berge SV, Aarsaether N, Littlewood-Evans A et al. Mural cell associated VEGF is required for organotypic vessel formation. PLoS ONE 2009; 4: e5798–e5808.

    Article  PubMed  PubMed Central  Google Scholar 

  14. Thorve VS, Kshirsagar AD, Vyawahare NS, Joshi VS, Ingale KG, Mohite RJ . Diabetes-induced erectile dysfunction: epidemiology, pathophysiology and management. J Diabetes Complications 2011; 25: 129–136.

    Article  PubMed  Google Scholar 

  15. Jin HR, Kim WJ, Song JS, Piao S, Tumurbaatar M, Shin SH et al. Intracavernous delivery of synthetic angiopoietin-1 protein as a novel therapeutic strategy for erectile dysfunction in the type II diabetic db/db mouse. J Sex Med 2010; 7: 3635–3646.

    Article  CAS  PubMed  Google Scholar 

  16. Podlasek CA, Zelner DJ, Harris JD, Meroz CL, Tang Y, McKenna KE et al. Altered Sonic hedgehog signaling is associated with morphological abnormalities in the penis of the BB/WOR diabetic rat. Biol Reprod 2003; 69: 816–827.

    Article  CAS  PubMed  Google Scholar 

  17. Albersen M, Lin G, Fandel TM, Zhang H, Qiu X, Lin CS et al. Functional, metabolic, and morphologic characteristics of a novel rat model of type 2 diabetes-associated erectile dysfunction. Urology 2011; 78: 476.e1–8.

    Article  Google Scholar 

  18. Cagliero E, Maiello M, Boeri D, Roy S, Lorenzi M . Increased expression of basement membrane components in human endothelial cells cultured in high glucose. J Clin Invest 1988; 82: 735–738.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  19. Hayden MR, Sowers JR, Tyagi SC . The central role of vascular extracellular matrix and basement membrane remodeling in metabolic syndrome and type 2 diabetes: the matrix preloaded. Cardiovasc Diabetol 2005; 4: 9–28.

    Article  PubMed  PubMed Central  Google Scholar 

  20. Britland ST, Young RJ, Sharma AK, Clarke BF . Relationship of endoneurial capillary abnormalities to type and severity of diabetic polyneuropathy. Diabetes 1990; 39: 909–913.

    Article  CAS  PubMed  Google Scholar 

Download references

Acknowledgements

This work was supported by grants from the Arthur Rock Foundation and the National Institutes of Health (DK045370).

Author information

Authors and Affiliations

  1. Department of Urology, Knuppe Molecular Urology Laboratory, School of Medicine, University of California, San Francisco, CA, USA

    H Ning, G Lin, T F Lue & C-S Lin

Authors
  1. H Ning
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. G Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. T F Lue
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. C-S Lin
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to C-S Lin.

Ethics declarations

Competing interests

The authors declare no conflict of interest.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ning, H., Lin, G., Lue, T. et al. A coculture system of cavernous endothelial and smooth muscle cells. Int J Impot Res 25, 63–68 (2013). https://doi.org/10.1038/ijir.2012.36

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1038/ijir.2012.36

Keywords

This article is cited by

Search

Advanced search

Quick links