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Title: | Pentosan polysulfate binds to STRO-1+ mesenchymal progenitor cells, is internalized, and modifies gene expression: A novel approach of pre-programing stem cells for therapeutic application requiring their chondrogenesis. | Authors: | Wu J.;Ghosh P.;Zannettino A.C.W.;Gronthos S.;Goldschlager T. ;Daly C.;Paton S.;Shimmon S. | Institution: | (Wu, Shimmon, Ghosh) Proteobioactives Pty. Ltd., PO Box 174, Balgowlah, Sydney, NSW 2093, Australia (Paton, Zannettino) Myeloma Research Laboratory, Faculty of Health and Medical Sciences, University of Adelaide and the Cancer Theme, Adelaide, SA 5000, Australia (Daly, Goldschlager) Department of Surgery, Monash University, Clayton, VIC 3168, Australia (Daly, Goldschlager) Department of Neurosurgery, Monash Medical Centre, Clayton, VIC 3168, Australia (Daly, Goldschlager, Ghosh) Ritchie Centre, Hudson Institute of Medical Research, Monash University, Clayton, VIC 3168, Australia (Gronthos) Mesenchymal Stem Cell Laboratory, Adelaide Medical School, University of Adelaide, Adelaide, SA 5000, Australia (Wu) Present Address: Minomic International Ltd, 75 Talavera Rd, Macquarie Park, NSW 2113, Australia (Shimmon) School of Mathematical and Physical Sciences, Faculty of Science, University of Technology Sydney, PO Box 123, Sydney, NSW 2007, Australia | Issue Date: | 23-Dec-2017 | Copyright year: | 2017 | Publisher: | BioMed Central Ltd. (E-mail: info@biomedcentral.com) | Place of publication: | United Kingdom | Publication information: | Stem Cell Research and Therapy. 8 (1) (no pagination), 2017. Article Number: 278. Date of Publication: 13 Dec 2017. | Journal: | Stem Cell Research and Therapy | Abstract: | Background: The pharmaceutical agent pentosan polysulfate (PPS) is known to induce proliferation and chondrogenesis of mesenchymal progenitor cells (MPCs) in vitro and in vivo. However, the mechanism(s) of action of PPS in mediating these effects remains unresolved. In the present report we address this issue by investigating the binding and uptake of PPS by MPCs and monitoring gene expression and proteoglycan biosynthesis before and after the cells had been exposed to limited concentrations of PPS and then re-established in culture in the absence of the drug (MPC priming). Method(s): Immuno-selected STRO-1+ mesenchymal progenitor stem cells (MPCs) were prepared from human bone marrow aspirates and established in culture. The kinetics of uptake, shedding, and internalization of PPS by MPCs was determined by monitoring the concentration-dependent loss of PPS media concentrations using an enzyme-linked immunosorbent assay (ELISA) and the uptake of fluorescein isothiocyanate (FITC)-labelled PPS by MPCs. The proliferation of MPCs, following pre-incubation and removal of PPS (priming), was assessed using the Wst-8 assay method, and proteoglycan synthesis was determined by the incorporation of 35SO4 into their sulphated glycosaminoglycans. The changes in expression of MPC-related cell surface antigens of non-primed and PPS-primed MPCs from three donors was determined using flow cytometry. RNA sequencing of RNA isolated from non-primed and PPS-primed MPCs from the same donors was undertaken to identify the genes altered by the PPS priming protocol. Result(s): The kinetic studies indicated that, in culture, PPS rapidly binds to MPC surface receptors, followed by internalisation and localization within the nucleus of the cells. Following PPS-priming of MPCs and a further 48 h of culture, both cell proliferation and proteoglycan synthesis were enhanced. Reduced expression of MPC-related cell surface antigen expression was promoted by the PPS priming, and RNA sequencing analysis revealed changes in the expression of 42 genes. Conclusion(s): This study has shown that priming of MPCs with low concentrations of PPS enhanced chondrogenesis and MPC proliferation by modifying their characteristic basal gene and protein expression. These findings offer a novel approach to re-programming mesenchymal stem cells for clinical indications which require the repair or regeneration of cartilaginous tissues such as in osteoarthritis and degenerative disc disease.Copyright © 2017 The Author(s). | DOI: | http://monash.idm.oclc.org/login?url=http://dx.doi.org/10.1186/s13287-017-0723-y | Link to associated publication: | Click here for full text options | PubMed URL: | 29237492 [http://www.ncbi.nlm.nih.gov/pubmed/?term=29237492] | ISSN: | 1757-6512 (electronic) | URI: | https://repository.monashhealth.org/monashhealthjspui/handle/1/38453 | Type: | Article | Subjects: | *pentosan polysulfate/pd [Pharmacology] stem cell culture sulfate RNA/ec [Endogenous Compound] proteoglycan/ec [Endogenous Compound] controlled study donor drug structure enzyme linked immunosorbent assay flow cytometry gene *gene expression regulation gene identification human human cell incubation time *internalization *mesenchymal stem cell *nuclear reprogramming priority journal protein expression proteoglycan synthesis receptor binding RNA sequence unclassified drug acta2 gene ADAMTSL4 gene ANK1 gene COL11A1 gene COL5A3 gene COMP gene DACT1 gene Enpp1 gene FLG gene GREM2 gene Hspb7 gene Large gene LMOD1 gene loxl4 gene LRRC15 gene MRVI1 gene scube3 gene SVL gene SYNPO2 gene TM4SF1 gene *stro1 protein/ec [Endogenous Compound] *pentosan polysulfate/an [Drug Analysis] membrane antigen/ec [Endogenous Compound] glycosaminoglycan/ec [Endogenous Compound] fluorescein isothiocyanate cell surface receptor/ec [Endogenous Compound] *cell marker/ec [Endogenous Compound] antigen expression article binding kinetics bone marrow biopsy cell assay cell culture cell labeling cell nucleus cell proliferation cell selection cellular distribution *chondrogenesis concentration response receptor binding priority journal protein expression proteoglycan synthesis RNA sequence stem cell culture antigen expression Article binding kinetics bone marrow biopsy cell assay cell culture cell labeling cell nucleus cell proliferation cell selection cellular distribution *chondrogenesis concentration response controlled study donor drug structure enzyme linked immunosorbent assay flow cytometry gene *gene expression regulation gene identification human human cell incubation time *internalization *mesenchymal stem cell *nuclear reprogramming |
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