Decellularized kidney in the presence of chondroitin sulfate as a natural 3D scaffold for stem cells

Document Type: Original Article


1 Department of Biology, Faculty of Sciences, Mashhad Branch, Islamic Azad University, Mashhad, Iran

2 Department of Biology, Faculty of Sciences, Ferdowsi University of Mashhad, Mashhad, Iran


Objective(s): Use of biological scaffolds and automating the cells directing process with materials such as growth factors and glycosaminoglycans (GAGs) in a certain path may have beneficial effects in tissue engineering and regenerative medicine in future. In this research, chondroitin sulfate sodium was used for impregnation of the scaffolds. It is a critical component in extracellular matrix and plays an important role in signaling pathway; however, little is known about its role within mammalian development and cell linage specification.
Materials and Methods: Due to its porous and appropriate structure and for putting cells in 3D space, the kidney of BALB/c mouse was selected and decellulalized using physical and chemical methods. After decellularization, the scaffold was impregnated in chondroitin sulfate solution (CS) for 24 hr. Then, 60×10 5 human adipose-derived mesenchymal stem cells were seeded on the scaffold to assess their behavior on day 5, 10, 15, 20, and 25.
Result: After 48 hr, DAPI staining approved completed decellularized kidney by 1% SDS (sodium dodecyl sulfate). Migration and establishment of a number of cells to the remaining area of the glomerulus was observed. In addition, cell accumulation on the scaffold surface as well as cells migration to the depth of kidney formed an epithelium-like structure. Up to the day 15, microscopic study of different days of seeding showed the gradual adhesion of large number of cells to the scaffold.
Conclusion: Glycosaminoglycan could be a right option for impregnation. It is used for smartification and strengthening of natural scaffolds and induction of some behaviors in stem cells.


1. Badylak SF. Regenerative medicine and developmental biology: the role of the extracellular matrix. Anat Rec B New Anat 2005; 287:36-41.

2. Badylak SF, Freytes DO, Gilbert TW. Extracellular matrix as a biological scaffold material: Structure and function. Acta Biomaterialia 2009; 5:1-13.

3. Nakayama KH, Batchelder CA, Lee CI, Tarantal AF. Decellularized rhesus monkey kidney as a three-dimensional scaffold for renal tissue engineering. Tissue Engineering Part A 2010; 16:2207-16.

4. Mardani M, Hashemibeni B, Ansar MM, Zarkesh Esfahani SH, Kazemi M, Goharian V, et al. Comparison between chondrogenic markers of differentiated chondrocytes from adipose derived stem cells and articular chondrocytes in vitro. Iran J Basic Med Sci 2013; 16:763-73.

5. Strachan LR, Condic ML. Neural crest motility and integrin regulation are distinct in cranial and trunk populations. Developmental Biology 2003; 259:288-302.

6. Rozario T, DeSimone DW. The extracellular matrix in development and morphogenesis: a dynamic view. Developmental Biology 2010; 341:126-140.

7. Linton JM, Martin GR, Reichardt LF. The ECM protein nephronectin promotes kidney development via integrin alpha8beta1-mediated stimulation of Gdnf expression. Development 2007; 134:2501-2509.

8. Neu R, Adams S, Munz B. Differential expression of entactin-1/nidogen-1 and entactin-2/nidogen-2 in myogenic differentiation. Differentiation 2006; 74:573-582.

9. Guilak F, Cohen DM, Estes BT, Gimble JM, Liedtke W, Chen CS. Control of stem cell fate by physical interactions with the extracellular matrix. Cell Stem Cell 2009 Jul 2; 5:17-26.

10. Zuk PA, Zhu M, Ashjian P, De Ugarte DA, Huang JI, Mizuno H, et al. Human adipose tissue is a source of multipotent stem cells. Molecular Biology Of The Cell 2002; 13:4279-4295.

11. Casteilla L, Planat-Benard V, Laharrague P, Cousin B. Adipose-derived stromal cells: Their identity and uses in clinical trials, an update. World J Stem Cells 2011; 3:25-33.

12. Lindroos B, Suuronen R, Miettinen S. The potential of adipose stem cells in regenerative medicine. Stem Cell Rev 2011; 7:269-291.

13. Mohammadzadeh E, Nikravesh MR, Jalali M, Fazel A, Ebrahimi V, Ebrahimzadeh-Bideskan AR. Immunohistochemical study of type III collagen expression during pre and post-natal rat skin morphogenesis. Iran J Basic Med Sci 2014; 17:196-200.

14. Hodde JP, Badylak SF, Brightman AO, Voytik-Harbin SL. Glycosaminoglycan content of small intestinal submucosa: a bioscaffold for tissue replacement. Tissue Eng 1996; 2:209-217.

15. Prinz RD, Willis CM, van Kuppevelt TH, Kluppel M. Biphasic role of chondroitin sulfate in cardiac differentiation of embryonic stem cells through inhibition of Wnt/beta-catenin signaling. PLoS One 2014; 9:e92381.

16. Crapo PM, Gilbert TW, Badylak SF. An overview of tissue and whole organ decellularization processes. Biomaterials 2011; 32:3233-3243.

17. Naderi S, Khayat Zadeh J, Mahdavi Shahri N, Nejad Shahrokh Abady K, Cheravi M, Baharara J,            et al. Three-dimensional scaffold from decellularized human gingiva for cell cultures: glycoconjugates and cell behavior. Cell J 2013; 15: 166-175.

18. Gilbert TW, Sellaro TL, Badylak SF. Decellularization of tissues and organs. Biomaterials 2006; 27:3675-3683.

19. Mahdavi Shahri N, Baharara J, Takbiri M, Khajeh Ahmadi S. In vitro decellularization of rabbit lung tissue. Cell J 2013; 15:83-88.

20. Haddad-Mashadrizeh A, Bahrami AR, Matin MM, Edalatmanesh MA, Zomorodipour A, Gardaneh M, et al. Human adipose-derived mesenchymal stem cells can survive and integrate into the adult rat eye following xenotransplantation. Xenotransplantation 2013; 20:165-176.

21. Badylak SF. The extracellular matrix as a scaffold for tissue reconstruction. Semin Cell  Dev Biol 2002; 13:377-383.

22. Ross EA, Williams MJ, Hamazaki T, Terada N, Clapp WL, Adin C, et al. Embryonic stem cells proliferate and differentiate when seeded into kidney scaffolds. J Am Soc of Nephrol 2009 ; 20:2338-2347.

23. Nakayama KH, Batchelder CA, Lee CI, Tarantal AF. Renal tissue engineering with decellularized rhesus monkey kidneys: age-related differences. Tissue Engin Part A 2011; 17:2891-2901.

24. Cebotari S, Tudorache I, Jaekel T, Hilfiker A, Dorfman S, Ternes W, et al. Detergent decellularization of heart valves for tissue engineering: toxicological effects of residual detergents on human endothelial cells. Artif Organs 2010; 34:206-210.

25. Sullivan DC, Mirmalek-Sani SH, Deegan DB, Baptista PM, Aboushwareb T, Atala A, et al. Decellularization methods of porcine kidneys for whole organ engineering using a high-throughput system. Biomaterials 2012;33:7756-7764.

26. Liu CX, Liu SR, Xu AB, Kang YZ, Zheng SB, Li HL, et al. Preparation of whole-kidney acellular matrix in rats by perfusion. Nan Fang Yi Ke Da Xue Xue Bao 2009; 29:979-982.

27. Walker PD. Alterations in renal tubular extracellular matrix components after ischemia-reperfusion injury to the kidney. Lab Investig 1994; 70:339-345.

28. Li HY, Liao CY, Lee KH, Chang HC, Chen YJ, Chao KC, et al. Collagen IV significantly enhances migration and transplantation of embryonic stem cells: involvement of alpha2beta1 integrin-mediated actin remodeling. Cell Transplant 2011; 20:893-907.

29. Vorotnikova E, McIntosh D, Dewilde A, Zhang J, Reing JE, Zhang L, et al. Extracellular matrix-derived products modulate endothelial and progenitor cell migration and proliferation in vitro and stimulate regenerative healing in vivo. Matrix Biol 2010; 29:690-700.

30. Kleinman HK, Klebe RJ, Martin GR. Role of collagenous matrices in the adhesion and growth of cells. J Cell Biol 1981; 88:473-485.

31. Michelini M, Franceschini V, Sihui Chen S,  Papini S, Rosellini A, Ciani F, et al. Primate embryonic stem cells create their own niche while differentiating in three-dimensional culture systems. Cell Prolifer 2006; 39:217-229.

32. Gerecht-Nir S, Ziskind A, Cohen S, Itskovitz-Eldor J. Human embryonic stem cells as an in vitro model for human vascular development and the induction of vascular differentiation. Lab investig 2003; 83:1811-1820.

33. Baharvand H, Hashemi SM, Kazemi Ashtiani S, Farrokhi A. Differentiation of human embryonic stem cells into hepatocytes in 2D and 3D culture systems in vitro. Int J Dev Biol 2006; 50:645-652.

34. Wight TN, Kinsella MG, Qwarnstrom EE. The role of proteoglycans in cell adhesion, migration and proliferation. Curr Opin Cell Biol 1992; 4:793-801.

35. Brendan AC, Lorna J. In vivo and in vitro applications of collagen-GAG scaffolds. Chem Engin J 2008; 137:102-121.

36. Huang KF, Hsu WC, Chiu WT, Wang JY. Functional improvement and neurogenesis after collagen-GAG matrix implantation into surgical brain trauma. Biomaterials 2012; 33:2067-2075.

37. Tierney CM, Jaasma MJ, O'Brien FJ. Osteoblast activity on collagen-GAG scaffolds is affected by collagen and GAG concentrations. J Biomed Mater Res A 2009; 91:92-101.

38. Vickers SM, Squitieri LS, Spector M. Effects of cross-linking type II collagen-GAG scaffolds on chondrogenesis in vitro: dynamic pore reduction promotes cartilage formation. Tissue Engin 2006; 12:1345-1355.

39. Frisch SM, Francis H. Disruption of epithelial cell-matrix interactions induces apoptosis. J Cell Biol 1994; 124:619-626.

40. Sugiyama H, Kashihara N, Maeshima Y, Okamoto K, Kanao K, Sekikawa T, et al. Regulation of survival and death of mesangial cells by extracellular matrix. Kidney Int 1998; 54:1188-1196.

41. Makino H, Sugiyama H, Kashihara N. Apoptosis and extracellular matrix-cell interactions in kidney disease. Kidney Int 2000; 77:S67-75.

42. Kessenbrock K, Plaks V, Werb Z. Matrix metalloproteinases: regulators of the tumor microenvironment. Cell 2010; 141:52-67.

43. Friedl P. Prespecification and plasticity: shifting mechanisms of cell migration. Curr Opin Cell Biol 2004; 16:14-23.