Clinical and Experimental Medicine

, Volume 15, Issue 4, pp 501–509 | Cite as

PDX-1 mRNA-induced reprogramming of mouse pancreas-derived mesenchymal stem cells into insulin-producing cells in vitro

  • Xing Rong Guo
  • Xiao Li Wang
  • Man Chol Li
  • Ya Hong Yuan
  • Yun Chen
  • Dan Dan Zou
  • Liu Jiao BianEmail author
  • Dong Sheng LiEmail author
Original Article


Pancreatic islet transplantation has remained an effective therapy for type 1 diabetes since 2000. Its widespread use has been prohibited by the shortage of suitable donors. It is critical to explore an applicable alternative for β-cell replacement. This study was performed to generate insulin-producing cells (IPCs) from pancreas-derived mesenchymal stem cells (pMSCs). pMSCs were isolated from discarded pancreatic tissue in the filter liquor during islet isolation procedure in mice and ex vivo expanded in culture. IPCs were induced by transfection of pancreas and duodenal transcription factor 1 (PDX-1) mRNA in vitro. Some islet characteristics were identified on pMSC-derived IPCs in mRNA and protein levels. Our results demonstrated that mouse pMSCs can be transdifferentiated into effective glucose-responsive insulin-producing cells through transfecting synthetic modified PDX-1 mRNA in vitro. The study of PDX-1 mRNA-induced pMSC reprogramming may pave the way toward the development of a novel β-cell source for the treatment of diabetes.


Pancreas-derived mesenchymal stem cells (pMSCs) Reprogramming PDX-1 mRNA Insulin-producing cells (IPCs) 



This study was financially supported by the Major Science and Technology Projects of China (No. 2013ZX10001-004-002-005), National Natural Science Foundation of China (No. 21075097), the National Natural Science Foundation of China (81070614), National Natural Science Foundation of Hubei province (2012FFA037), Natural Science Support Foundation for graduated students of Hubei University of Medicine (2011QDZR-8) and Foundation of Hubei Educational Committee(B20122411). Authors are grateful to Dr. Long-Jun Dai of University of British Columbia in Canada for his critical review and discussion of the manuscript.

Conflict of interest


Ethical standards

All animal studies have been approved by the appropriate ethics committee and have therefore been performed in accordance with the ethical standards.

Supplementary material

10238_2014_319_MOESM1_ESM.tif (1.8 mb)
Supplementary Material Figure 1. A. Immunofluorescence analyses of PDX-1 expression in islets and islet-derived MSCs. Fresh islets cultured for 8 days in modified RPMI 1640 media containing 20 ng/ml bFGF and EGF showing outgrowth of the monolayer of cells—islet-derived MSCs. PDX-1 specially expressed in islets, but undetected in islet-derived MSCs, which is consistent with the pMSCs obtained from our method. (black↑ for Islets, red↑ for pMSCs) B. GFP mRNA transfected into pMSCs as a control. The expression of GFP was determined with FACS (a) and examined under a fluorescent microscope (b). C. Morphological shape of differentiated pMSCs into IPCs in the first (a) and second approaches (b) on 8 days. (TIFF 1839 kb)
10238_2014_319_MOESM2_ESM.tif (2.2 mb)
Supplementary Material Figure 2. The primary culture of the isolated cells contained a heterogeneous cell population with both round and fibroblast-like cells. Round cells grew fast and fibroblast-like cells hardly grew on day 3(A); fibroblast-like cells began to grow, round cells stop developing on day 6(B); fibroblast-like cells proliferated very well on day 10(C). (black↑for round cells, red↑for pMSCs) (TIFF 2298 kb)
10238_2014_319_MOESM3_ESM.pdf (123 kb)
Supplementary Material Table 1. Primer sequences and PCR product size (PDF 122 kb)


  1. 1.
    Lebastchi J, Deng S, Lebastchi AH, et al. Immune therapy and beta-cell death in type 1 diabetes. Diabetes. 2013;62(5):1676–80.CrossRefPubMedPubMedCentralGoogle Scholar
  2. 2.
    Hematti P, Kim J, Stein AP, et al. Potential role of mesenchymal stromal cells in pancreatic islet transplantation. Transplant Rev. 2013;27(1):21–9.CrossRefGoogle Scholar
  3. 3.
    Gershengorn MC, Geras-Raaka E, Hardikar AA, et al. Extra views are better islet cell precursors generated by epithelial-to-mesenchymal transition? Cell Cycle. 2005;4(3):380–2.CrossRefPubMedGoogle Scholar
  4. 4.
    Eberhardt M, Salmon P, von Mach M-A, et al. Multipotential nestin and Isl-1 positive mesenchymal stem cells isolated from human pancreatic islets. Biochem Biophys Res commun. 2006;345(3):1167–76.CrossRefPubMedGoogle Scholar
  5. 5.
    Atouf F, Park CH, Pechhold K, et al. No evidence for mouse pancreatic beta-cell epithelial-mesenchymal transition in vitro. Diabetes. 2007;56(3):699–702.CrossRefPubMedGoogle Scholar
  6. 6.
    Chase LG, Ulloa-Montoya F, Kidder BL, et al. Islet-derived fibroblast-like cells are not derived via epithelial-mesenchymal transition from Pdx-1 or insulin-positive cells. Diabetes. 2007;56(1):3–7.CrossRefPubMedGoogle Scholar
  7. 7.
    Gallo R, Gambelli F, Gava B, et al. Generation and expansion of multipotent mesenchymal progenitor cells from cultured human pancreatic islets. Cell Death Differ. 2007;14(11):1860–71.CrossRefPubMedGoogle Scholar
  8. 8.
    Davani B, Ikonomou L, Raaka BM, et al. Human islet-derived precursor cells are mesenchymal stromal cells that differentiate and mature to hormone-expressing cells in vivo. Stem Cells. 2007;25(12):3215–22.CrossRefPubMedGoogle Scholar
  9. 9.
    Kim SJ, Choi YS, Ko ES, et al. Glucose-stimulated insulin secretion of various mesenchymal stem cells after insulin-producing cell differentiation. J Biosci Bioeng. 2012;113(6):771–7.CrossRefPubMedGoogle Scholar
  10. 10.
    Anzalone R, Iacono ML, Loria T, et al. Wharton’s jelly mesenchymal stem cells as candidates for beta cells regeneration: extending the differentiative and immunomodulatory benefits of adult mesenchymal stem cells for the treatment of type 1 diabetes. Stem Cell Rev Rep. 2011;7(2):342–63.CrossRefGoogle Scholar
  11. 11.
    Zulewski H, Abraham EJ, Gerlach MJ, et al. Multipotential nestin-positive stem cells isolated from adult pancreatic islets differentiate ex vivo into pancreatic endocrine, exocrine, and hepatic phenotypes. Diabetes. 2001;50(3):521–33.CrossRefPubMedGoogle Scholar
  12. 12.
    Zanini C, Bruno S, Mandili G, et al. Differentiation of mesenchymal stem cells derived from pancreatic islets and bone marrow into islet-like cell phenotype. PLoS ONE. 2011;6(12):e28175.CrossRefPubMedPubMedCentralGoogle Scholar
  13. 13.
    Cerf ME. Transcription factors regulating ?2-cell function. Eur J Endocrinol. 2006;155(5):671–9.CrossRefPubMedGoogle Scholar
  14. 14.
    Jafarian A, Taghikhani M, Abroun S, et al. Generation of high-yield insulin producing cells from human bone marrow mesenchymal stem cells. Mol Biol Rep. 2014;41(7):4783–94.CrossRefPubMedGoogle Scholar
  15. 15.
    Wang H, Yang Y, Ho G, et al. Programming of human umbilical cord mesenchymal stem cells in vitro to promote pancreatic gene expression. Mol Med Rep. 2013;8(3):769–74.PubMedGoogle Scholar
  16. 16.
    Yuan H, Li J, Xin N, et al. Expression of Pdx1 mediates differentiation from mesenchymal stem cells into insulin-producing cells. Mol Biol Rep. 2010;37(8):4023–31.CrossRefPubMedGoogle Scholar
  17. 17.
    Leonhardt C, Schwake G, Stogbauer TR, et al. Single-cell mRNA transfection studies: delivery, kinetics and statistics by numbers. Nanomed Nanotechnol Biol Med. 2014;10(4):679–88.CrossRefGoogle Scholar
  18. 18.
    Li DS, Yuan YH, Tu HJ, et al. A protocol for islet isolation from mouse pancreas. Nat Protoc. 2009;4(11):1649–52.CrossRefPubMedGoogle Scholar
  19. 19.
    Wang XL, Hu P, Guo XR, et al. Reprogramming Human Umbilical Cord Mesenchymal Stem Cells to Islet-like Cells with in vitro synthesized PDX1 mRNA. Cytotherapy. 2014;16(11):1519–27.Google Scholar
  20. 20.
    Tropepe V, Sibilia M, Ciruna BG, et al. Distinct neural stem cells proliferate in response to EGF and FGF in the developing mouse telencephalon. Dev Biol. 1999;208(1):166–88.CrossRefPubMedGoogle Scholar
  21. 21.
    Otonkoski T, Beattie G, Mally M, et al. Nicotinamide is a potent inducer of endocrine differentiation in cultured human fetal pancreatic cells. J Clin Invest. 1993;92(3):1459.CrossRefPubMedPubMedCentralGoogle Scholar
  22. 22.
    Zalzman M, Anker-Kitai L, Efrat S. Differentiation of human liver-derived, insulin-producing cells toward the β-cell phenotype. Diabetes. 2005;54(9):2568–75.CrossRefPubMedGoogle Scholar
  23. 23.
    Oh SH, Muzzonigro TM, Bae SH, et al. Adult bone marrow-derived cells trans-differentiating into insulin-producing cells for the treatment of type I diabetes. Lab Invest. 2004;84(5):607–17.CrossRefPubMedGoogle Scholar
  24. 24.
    Dominici M, Le Blanc K, Mueller I, et al. Minimal criteria for defining multipotent mesenchymal stromal cells. The international society for cellular therapy position statement. Cytotherapy. 2006;8(4):315–7.CrossRefPubMedGoogle Scholar
  25. 25.
    Moniri MR, Sun XY, Rayat J, et al. TRAIL-engineered pancreas-derived mesenchymal stem cells: characterization and cytotoxic effects on pancreatic cancer cells. Cancer Gene Ther. 2012;19(9):652–8.CrossRefPubMedGoogle Scholar
  26. 26.
    Limbert C, Jakob F, Ebert R, et al. Human pancreatic islet-derived precursor cells display mesenchymal stem cell features and differentiation capacity. J Stem Cells Regen Med. 2007;2(1):93–4.PubMedGoogle Scholar
  27. 27.
    Gu G, Dubauskaite J, Melton DA. Direct evidence for the pancreatic lineage: NGN3+ cells are islet progenitors and are distinct from duct progenitors. Development. 2002;129(10):2447–57.PubMedGoogle Scholar
  28. 28.
    Warren L, Manos PD, Ahfeldt T, et al. Highly efficient reprogramming to pluripotency and directed differentiation of human cells with synthetic modified mRNA. Cell Stem Cell. 2010;7(5):618–30.CrossRefPubMedPubMedCentralGoogle Scholar
  29. 29.
    Yakubov E, Rechavi G, Rozenblatt S, et al. Reprogramming of human fibroblasts to pluripotent stem cells using mRNA of four transcription factors. Biochem Biophys Res Commun. 2010;394(1):189–93.CrossRefPubMedGoogle Scholar
  30. 30.
    Gershengorn MC, Hardikar AA, Wei C, et al. Epithelial-to-mesenchymal transition generates proliferative human islet precursor cells. Science. 2004;306(5705):2261–4.CrossRefPubMedGoogle Scholar
  31. 31.
    Gao T, McKenna B, Li C, et al. Pdx1 maintains beta cell identity and function by repressing an alpha cell program. Cell Metab. 2014;19(2):259–71.CrossRefPubMedPubMedCentralGoogle Scholar
  32. 32.
    Schwitzgebel VM, Mamin A, Brun T, et al. Agenesis of human pancreas due to decreased half-life of insulin promoter factor 1. J Clin Endocrinol Metab. 2003;88(9):4398–406.CrossRefPubMedGoogle Scholar
  33. 33.
    Li HT, Jiang FX, Shi P, et al. In vitro reprogramming of rat bone marrow-derived mesenchymal stem cells into insulin-producing cells by genetically manipulating negative and positive regulators. Biochem Biophys Res Commun. 2012;420(4):793–8.CrossRefPubMedGoogle Scholar
  34. 34.
    Zhou Q, Brown J, Kanarek A, et al. In vivo reprogramming of adult pancreatic exocrine cells to &bgr;-cells. Nature. 2008;455(7213):627–32.CrossRefPubMedGoogle Scholar
  35. 35.
    Chun SY, Mack DL, Moorefield E, et al. Pdx1 and controlled culture conditions induced differentiation of human amniotic fluid-derived stem cells to insulin-producing clusters. J Tissue Eng Regen Med. 2012;13(10):1631–49.Google Scholar
  36. 36.
    Raikwar SP, Zavazava N. PDX1-engineered embryonic stem cell-derived insulin producing cells regulate hyperglycemia in diabetic mice. Transplant Res. 2012;1(1):19.CrossRefPubMedPubMedCentralGoogle Scholar
  37. 37.
    Jemielity J, Stepinski J, Jaremko M, et al. Synthesis of novel mRNA 5′ cap-analogues: dinucleoside P1, P3-tri-, P1, P4-tetra-, and P1, P5-pentaphosphates. Nucleosides, Nucleotides Nucleic Acids. 2003;22(5–8):691–4.CrossRefPubMedGoogle Scholar
  38. 38.
    Stepinski J, Waddell C, Stolarski R, et al. Synthesis and properties of mRNAs containing the novel “anti-reverse” cap analogs 7-methyl(3′-O-methyl)GpppG and 7-methyl (3′-deoxy)GpppG. RNA. 2001;7(10):1486–95.PubMedPubMedCentralGoogle Scholar
  39. 39.
    Bangel-Ruland N, Tomczak K, Fernandez Fernandez E, et al. Cystic fibrosis transmembrane conductance regulator-mRNA delivery: a novel alternative for cystic fibrosis gene therapy. J Gene Med. 2013;15(11–12):414–26.CrossRefPubMedGoogle Scholar
  40. 40.
    Bar-Nur O, Russ HA, Efrat S, et al. Epigenetic memory and preferential lineage-specific differentiation in induced pluripotent stem cells derived from human pancreatic islet beta cells. Cell Stem Cell. 2011;9(1):17–23.CrossRefPubMedGoogle Scholar
  41. 41.
    Lakey JR, Kneteman NM, Rajotte RV, et al. Effect of core pancreas temperature during cadaveric procurement on human islet isolation and functional viability. Transplantation. 2002;73(7):1106–10.CrossRefPubMedGoogle Scholar

Copyright information

© Springer-Verlag Italia 2014

Authors and Affiliations

  • Xing Rong Guo
    • 1
    • 2
  • Xiao Li Wang
    • 2
  • Man Chol Li
    • 2
  • Ya Hong Yuan
    • 2
  • Yun Chen
    • 2
  • Dan Dan Zou
    • 2
  • Liu Jiao Bian
    • 1
    Email author
  • Dong Sheng Li
    • 2
    Email author
  1. 1.College of Life SciencesNorthwest UniversityXi’anChina
  2. 2.Hubei Key Laboratory of Embryonic Stem Cell Research, Taihe HospitalHubei University of MedicineShiyanChina

Personalised recommendations