Compositional and structural analysis of glycosaminoglycans in cell-derived extracellular matrices
The extracellular matrix (ECM) is a highly dynamic and complex meshwork of proteins and glycosaminoglycans (GAGs) with a crucial role in tissue homeostasis and organization not only by defining tissue architecture and mechanical properties, but also by providing chemical cues that regulate major biological processes. GAGs are associated with important physiological functions, acting as modulators of signaling pathways regulating several cellular processes such as cell growth and differentiation. Recently, in vitro fabricated cell-derived ECM have emerged as promising materials for regenerative medicine due to their ability of better recapitulate the native ECM-like composition and structure, without the limitations of availability and pathogen transfer risks of tissue-derived ECM scaffolds. However, little is known about the molecular and more specifically, GAG composition of these cell-derived ECM. In this study, three different cell-derived ECM were produced in vitro and characterized in terms of their GAG content, composition and sulfation patterns using a highly sensitive liquid chromatography-tandem mass spectrometry technique. Distinct GAG compositions and disaccharide sulfation patterns were verified for the different cell-derived ECM. Additionally, the effect of decellularization method on the GAG and disaccharide relative composition was also assessed. In summary, the method presented here offers a novel approach to determine the GAG composition of cell-derived ECM, which we believe is critical for a better understanding of ECM role in directing cellular responses and has the potential for generating important knowledge to use in the development of novel ECM-like biomaterials for tissue engineering applications.
KeywordsGlycosaminoglycans Compositional analysis, Cell-derived extracellular matrix Disaccharides Chondrocytes Mesenchymal stem cells
This work was supported by funding received by iBB-Institute for Bioengineering and Biosciences through Programa Operacional Regional de Lisboa 2020 (Project N. 007317), through the EU COMPETE Program and from National Funds through FCT-Portuguese Foundation for Science and Technology under the Programme grant UID/BIO/04565/2013 and by the European Union Framework Programme for Research and Innovation HORIZON 2020, under the Teaming Grant agreement No 739572 – The Discoveries Centre for Regenerative and Precision Medicine. This study was also supported by Center for Biotechnology and Interdisciplinary Studies-Rensselaer Polytechnic Institute funds and by the National Institutes of Health (Grant # DK111958). João C. Silva and Marta S. Carvalho would also like to acknowledge FCT for financial support through the scholarships SFRH/BD/105771/2014 and SFRH/BD/52478/2014, respectively.
Compliance with ethical standards
Conflict of interest
The authors declare no conflict of interest.
This work does not contain any studies with human participants or animals performed by any of the authors.
- 1.Lu, H., Hoshiba, T., Kawazoe, N., Koda, I., Song, M., Chen, G.: Cultured cell-derived extracellular matrix scaffolds for tissue engineering. Biomaterials. 32, 9658–9666 (2011). https://doi.org/10.1016/j.biomaterials.2011.08.091 CrossRefGoogle Scholar
- 4.Gilbert, T.W., Sellaro, T.L., Badylak, S.F.: Decellularization of tissues and organs. Biomaterials. 27, 3675–3683 (2006). https://doi.org/10.1016/j.biomaterials.2006.02.014 Google Scholar
- 5.Badylak, S.F., Taylor, D., Uygun, K.: Whole organ tissue engineering: Decellularization and Recellularization of three-dimensional matrix scaffolds. Annu. Rev. Biomed. Eng. 13, 27–53 (2010). https://doi.org/10.1146/annurev-bioeng-071910-124743 CrossRefGoogle Scholar
- 8.Lu, H., Hoshiba, T., Kawazoe, N., Chen, G.: Autologous extracellular matrix scaffolds for tissue engineering. Biomaterials. 32, 2489–2499 (2011). https://doi.org/10.1016/j.biomaterials.2010.12.016 CrossRefGoogle Scholar
- 9.Kang, Y., Kim, S., Bishop, J., Khademhosseini, A., Yang, Y.: The osteogenic differentiation of human bone marrow MSCs on HUVEC-derived ECM and β-TCP scaffold. Biomaterials. 33, 6998–7007 (2012). https://doi.org/10.1016/j.biomaterials.2012.06.061 CrossRefGoogle Scholar
- 10.Zeitouni, S., Krause, U., Clough, B.H., Halderman, H., Falster, A., Blalock, D.T., Chaput, C.D., Sampson, H.W., Gregory, C.A.: Human mesenchymal stem cell-derived matrices for enhanced osteoregeneration. Sci. Transl. Med. 4, 132–155 (2012). https://doi.org/10.1126/scitranslmed.3003396 CrossRefGoogle Scholar
- 11.Yang, Y., Lin, H., Shen, H., Wang, B., Lei, G., Tuan, R.S.: Mesenchymal stem cell-derived extracellular matrix enhances chondrogenic phenotype of and cartilage formation by encapsulated chondrocytes in vitro and in vivo. Acta Biomater. 69, 71–82 (2018). https://doi.org/10.1016/j.actbio.2017.12.043 CrossRefGoogle Scholar
- 13.Dominici, M., Le Blanc, K., Mueller, I., Slaper-Cortenbach, I., Marini, F.C., Krause, D.S., Deans, R.J., Keating, A., Prockop, D.J., Horwitz, E.M.: Minimal criteria for defining multipotent mesenchymal stromal cells. The International Society for Cellular Therapy position statement. Cytotherapy. 8, 315–317 (2006). https://doi.org/10.1080/14653240600855905 CrossRefGoogle Scholar
- 14.Murphy, M.B., Moncivais, K., Caplan, A.I.: Mesenchymal stem cells: Environmentally responsive therapeutics for regenerative medicine, (2013)Google Scholar
- 16.Park, Y.B., Seo, S., Kim, J.A., Heo, J.C., Lim, Y.C., Ha, C.W.: Effect of chondrocyte-derived early extracellular matrix on chondrogenesis of placenta-derived mesenchymal stem cells. Biomed. Mater. 10, (2015). https://doi.org/10.1088/1748-6041/10/3/035014
- 22.Gasimli, L., Hickey, A.M., Yang, B., Li, G., Dela Rosa, M., Nairn, A.V., Kulik, M.J., Dordick, J.S., Moremen, K.W., Dalton, S., Linhardt, R.J.: Changes in glycosaminoglycan structure on differentiation of human embryonic stem cells towards mesoderm and endoderm lineages. Biochim. Biophys. Acta, Gen. Subj. 1840, 1993–2003 (2014). https://doi.org/10.1016/j.bbagen.2014.01.007 CrossRefGoogle Scholar
- 27.Dombrowski, C., Song, S.J., Chuan, P., Lim, X., Susanto, E., Sawyer, A.A., Woodruff, M.A., Hutmacher, D.W., Nurcombe, V., Cool, S.M.: Heparan sulfate mediates the proliferation and differentiation of rat mesenchymal stem cells. Stem Cells Dev. 18, 661–670 (2009). https://doi.org/10.1089/scd.2008.0157 CrossRefGoogle Scholar
- 28.Manton, K.J., Leong, D.F.M., Cool, S.M., Nurcombe, V.: Disruption of heparan and chondroitin sulfate signaling enhances mesenchymal stem cell-derived osteogenic differentiation via bone morphogenetic protein signaling pathways. Stem Cells. 25, 2845–2854 (2007). https://doi.org/10.1634/stemcells.2007-0065 CrossRefGoogle Scholar
- 30.Pfeifer, C.G., Berner, A., Koch, M., Krutsch, W., Kujat, R., Angele, P., Nerlich, M., Zellner, J.: Higher ratios of hyaluronic acid enhance chondrogenic differentiation of human MSCs in a hyaluronic acid-gelatin composite scaffold. Materials (Basel). 9, (2016). https://doi.org/10.3390/ma9050381
- 31.Christiansen-Weber, T., Noskov, A., Cardiff, D., Garitaonandia, I., Dillberger, A., Semechkin, A., Gonzalez, R., Kern, R.: Supplementation of specific carbohydrates results in enhanced deposition of chondrogenic-specific matrix during mesenchymal stem cell differentiation. J. Tissue Eng. Regen. Med. 12, 1261–1272 (2018). https://doi.org/10.1002/term.2658 CrossRefGoogle Scholar
- 32.Amann, E., Wolff, P., Breel, E., van Griensven, M., Balmayor, E.R.: Hyaluronic acid facilitates chondrogenesis and matrix deposition of human adipose derived mesenchymal stem cells and human chondrocytes co-cultures. Acta Biomater. 52, 130–144 (2017). https://doi.org/10.1016/j.actbio.2017.01.064 CrossRefGoogle Scholar
- 33.Weyers, A., Yang, B., Yoon, D.S., Park, J.-H., Zhang, F., Lee, K.B., Linhardt, R.J.: A structural analysis of Glycosaminoglycans from lethal and nonlethal breast Cancer tissues: toward a novel class of Theragnostics for personalized medicine in oncology? OMICS 16, 79–89 (2012). https://doi.org/10.1089/omi.2011.0102 CrossRefGoogle Scholar
- 34.Heiskanen, A., Hirvonen, T., Salo, H., Impola, U., Olonen, A., Laitinen, A., Tiitinen, S., Natunen, S., Aitio, O., Miller-Podraza, H., Wuhrer, M., Deelder, A.M., Natunen, J., Laine, J., Lehenkari, P., Saarinen, J., Satomaa, T., Valmu, L.: Glycomics of bone marrow-derived mesenchymal stem cells can be used to evaluate their cellular differentiation stage. Glycoconj. J. 26, 367–384 (2009). https://doi.org/10.1007/s10719-008-9217-6 CrossRefGoogle Scholar
- 37.Sun, X., Li, L., Overdier, K.H., Ammons, L.A., Douglas, I.S., Burlew, C.C., Zhang, F., Schmidt, E.P., Chi, L., Linhardt, R.J.: Analysis of Total human urinary glycosaminoglycan disaccharides by liquid chromatography-tandem mass spectrometry. Anal. Chem. 87, 6220–6227 (2015). https://doi.org/10.1021/acs.analchem.5b00913 CrossRefGoogle Scholar
- 38.Oguma, T., Tomatsu, S., Montano, A.M., Okazaki, O.: Analytical method for the determination of disaccharides derived from keratan, heparan, and dermatan sulfates in human serum and plasma by high-performance liquid chromatography/turbo ionspray ionization tandem mass spectrometry. Anal. Biochem. 368, 79–86 (2007). https://doi.org/10.1016/j.ab.2007.05.016 CrossRefGoogle Scholar
- 41.Liu, X., Krishnamoorthy, D., Lin, L., Xue, P., Zhang, F., Chi, L., Linhardt, R.J., Iatridis, J.C.: A method for characterising human intervertebral disc glycosaminoglycan disaccharides using liquid chromatography-mass spectrometry with multiple reaction monitoring. Eur. Cell. Mater. 35, 117–131 (2018). https://doi.org/10.22203/eCM.v035a09 CrossRefGoogle Scholar
- 42.Dos Santos, F., Andrade, P.Z., Boura, J.S., Abecasis, M.M., da Silva, C.L., Cabral, J.M.S.: Ex vivo expansion of human mesenchymal stem cells: a more effective cell proliferation kinetics and metabolism under hypoxia. J. Cell. Physiol. 223, 27–35 (2010). https://doi.org/10.1002/jcp.21987 Google Scholar
- 44.Guneta, V., Zhou, Z., Tan, N.S., Sugii, S., Wong, M.T.C., Choong, C.: Recellularization of decellularized adipose tissue-derived stem cells: role of the cell-secreted extracellular matrix in cellular differentiation. Biomater. Sci. 6, 168–178 (2018). https://doi.org/10.1039/c7bm00695k CrossRefGoogle Scholar
- 47.Ragelle, H., Naba, A., Larson, B.L., Zhou, F., Prijić, M., Whittaker, C.A., Del Rosario, A., Langer, R., Hynes, R.O., Anderson, D.G.: Comprehensive proteomic characterization of stem cell-derived extracellular matrices. Biomaterials. 128, 147–159 (2017). https://doi.org/10.1016/j.biomaterials.2017.03.008 CrossRefGoogle Scholar