Stem Cell Reviews and Reports

, Volume 6, Issue 4, pp 523–531 | Cite as

The Effect of Human Umbilical Cord Blood Cells on Survival and Cytokine Production by Post-Ischemic Astrocytes in Vitro

  • Lixian Jiang
  • Samuel Saporta
  • Ning Chen
  • Cyndy Davis Sanberg
  • Paul Sanberg
  • Alison WillingEmail author


Cerebral ischemia induces death of all neural cell types within the region affected by the loss of blood flow. We have shown that administering human umbilical cord blood cells after a middle cerebral artery occlusion in rats significantly reduces infarct size, presumably by rescuing cells within the penumbra. In this study we examined whether the cord blood cells enhanced astrocyte survival in an in vitro model of hypoxia with reduced glucose availability. Primary astrocyte cultures were incubated for 2 h in no oxygen (95% N, 5% CO2) and low glucose (1% compared to 4.5%) media. Cord blood mononuclear cells were added to half the cultures at the beginning of hypoxia. Astrocyte viability was determined using fluorescein diacetate/propidium iodide (FDA/PI) labeling and cytokine production by the astrocytes measured using ELISA. In some studies, T cells, B cells or monocytes/macrophages isolated from the cord blood mononuclear fraction with magnetic antibody cell sorting (MACS) were used instead to determine which cellular component of the cord blood mononuclear fraction was responsible for the observed effects. Co-culturing mononuclear cord blood cells with astrocytes during hypoxia stimulated production of IL-6 and IL-10 during hypoxia. The cord blood T cells decreased survival of the astrocytes after hypoxia but had no effect on the examined cytokines. Our data demonstrate that the tested cord blood fractions do not enhance astrocyte survival when delivered individually, suggesting there is either another cellular component that is neuroprotective or an interaction of all the cells is essential for protection.


Monocytes Macrophages T cell B cell Cytokines Hypoxia In vitro 



This study was funded in part by the American Heart Association (AEW, grants 0355183B & 0555266B) and the National Institutes of Health (AEW, R01 NS052839).

Conflict of Interest

Human umbilical cord blood cells were provided by Saneron CCEL Therapeutics, Inc. SS and AEW were consultants to Saneron CCEL Therapeutics, Inc. PRS is co-founder of Saneron CCEL Therapeutics, Inc. AEW and PRS are inventors on cord-blood related patents. CDS is Vice President for Research at Saneron CCEL Therapeutics.


  1. 1.
    Newcomb, J. D., Ajmo, C. T., Davis Sanberg, C., Sanberg, P. R., Pennypacker, K. R., & Willing, A. E. (2006). Timing of cord blood treatment after experimental stroke determines therapeutic efficacy. Cell Transplantation, 15, 213–223.CrossRefPubMedGoogle Scholar
  2. 2.
    Vendrame, M., Cassady, C. J., Newcomb, J., et al. (2004). Infusion of human umbilical cord blood cells in a rat model of stroke dose-dependently rescues behavioral deficits and reduces infarct volume. Stroke, 35, 2390–2395.CrossRefPubMedGoogle Scholar
  3. 3.
    Willing, A. E., Lixian, J., Milliken, M., et al. (2003). Intravenous versus intrastriatal cord blood administration in a rodent model of stroke. Journal of Neuroscience Research, 73, 296–307.CrossRefPubMedGoogle Scholar
  4. 4.
    Borlongan, C. V., Hadman, M., Davis Sanberg, C., & Sanberg, P. R. (2004). CNS entry of peripherally injected umbilical cord blood cells is not required for neuroprotection in stroke. Stroke, 35, 2385–2389.CrossRefPubMedGoogle Scholar
  5. 5.
    Vendrame, M., Gemma, C., de Mesquita, D., et al. (2005). Anti-inflammatory effects of human cord blood cells in a rat model of stroke. Stem Cells and Development, 14, 595–604.CrossRefPubMedGoogle Scholar
  6. 6.
    Hall, A. A., Leonardo, C. C., Collier, L. A., Rowe, D. D., Willing, A. E., & Pennypacker, K. R. (2009). Delayed treatments for stroke influence neuronal death in rat organotypic slice cultures subjected to oxygen glucose deprivation. Neuroscience, 164, 470–477.CrossRefPubMedGoogle Scholar
  7. 7.
    Hall, A., Guyer, A., Leonardo, C., et al. (2009). Umbilical cord blood cells directly suppress ischemic oligodendrocyte cell death. Journal of Neuroscience Research, 87, 333–341.CrossRefPubMedGoogle Scholar
  8. 8.
    Bracci-Laudiero, L., Celestino, D., Starace, G., et al. (2003). CD34-positive cells in human umbilical cord blood express nerve growth factor and its specific receptor TrkA. Journal of Neuroimmunology, 136, 130–139.CrossRefPubMedGoogle Scholar
  9. 9.
    Arien-Zakay, H., Lecht, S., Bercu, M. M., et al. (2009). Neuroprotection by cord blood neural progenitors involves antioxidants, neurotrophic and angiogenic factors. Experimental Neurology, 216, 83–94.CrossRefPubMedGoogle Scholar
  10. 10.
    Fan, C.-G., Zhang, Q.-J., Tang, F.-W., Han, Z.-B., Wang, G.-S., & Han, Z.-C. (2005). Human umbilical cord blood cells express neurotrophic factors. Neuroscience Letters, 380, 322–325.CrossRefPubMedGoogle Scholar
  11. 11.
    Zwart, I., Hill, A. J., Al-Allaf, F., et al. (2009). Umbilical cord blood mesenchymal stromal cells are neuroprotective and promote regeneration in a rat optic tract model. Experimental Neurology, 216, 439–448.CrossRefPubMedGoogle Scholar
  12. 12.
    Neuhoff, S., Moers, J., Rieks, M., et al. (2007). Proliferation, differentiation, and cytokine secretion of human umbilical cord blood–derived mononuclear cells in vitro. Experimental Hematology, 35, 1119–1131.CrossRefPubMedGoogle Scholar
  13. 13.
    McGuckin, C. P., Forraz, N., Allouard, Q., & Pettengell, R. (2004). Umbilical cord blood stem cells can expand hematopoietic and neuroglial progenitors in vitro. Experimental Cell Research, 295, 350–359.CrossRefPubMedGoogle Scholar
  14. 14.
    Hau, S., Reich, D. M., Scholz, M., et al. (2008). Evidence for neuroprotective properties of human umbilical cord blood cells after neuronal hypoxia in vitro. BMC Neuroscience, 9, 30.CrossRefPubMedGoogle Scholar
  15. 15.
    Jiang, L., Womble, T., Saporta, S., et al. (2010). Human umbilical cord blood cells depress the microglial inflammatory response in vitro. Stem Cells and Development, 19, 221–227.CrossRefPubMedGoogle Scholar
  16. 16.
    Ehrlich, L. C., Peterson, P. K., & Hu, S. (1999). Interleukin (IL)-1beta-mediated apoptosis of human astrocytes. NeuroReport, 10, 1849–1852.CrossRefPubMedGoogle Scholar
  17. 17.
    Suk, K., Lee, J., Hur, J., et al. (2001). Activation-induced cell death of rat astrocytes. Brain Research, 900, 342–347.CrossRefPubMedGoogle Scholar
  18. 18.
    Niu, F., Zhang, X., Chang, L., et al. (2009). Trichostatin A enhances OGD-astrocyte viability by inhibiting inflammatory reaction mediated by NF-κB. Brain Research Bulletin, 78, 342–346.CrossRefPubMedGoogle Scholar
  19. 19.
    Schultz, C., Reiss, I., Bucsky, P., et al. (2000). Maturational changes of lymphocyte surface antigens in human blood: comparison between fetuses, neonates and adults. Biology of the Neonate, 78, 77–82.CrossRefPubMedGoogle Scholar
  20. 20.
    Harris, D. P., Haynes, L., Sayles, P. C., et al. (2000). Reciprocal regulation of polarized cytokine production by effector B and T cells. Nature Immunology, 1, 475–482.CrossRefPubMedGoogle Scholar
  21. 21.
    Yu, P., Wang, Y., Chin, R. K., et al. (2002). B cells control the migration of a subset of dendritic cells into B cell follicles via CXC chemokine ligand 13 in a lymphotoxin-dependent fashion. Journal of Immunology, 168, 5117–5123.Google Scholar
  22. 22.
    Fogelstrand, L., Hulthe, J., Hulten, L. M., Wiklund, O., & Fagerberg, B. (2004). Monocytic expression of CD14 and CD18, circulating adhesion molecules and inflammatory markers in women with diabetes mellitus and impaired glucose tolerance. Diabetologia, 47, 1948–1952.CrossRefPubMedGoogle Scholar
  23. 23.
    Anderson, D. C., Schmalstieg, F. C., Kohl, S., et al. (1984). Abnormalities of polymorphonuclear leukocyte function associated with a heritable deficiency of a high molecular weight surface glycoprotein (gp138): common relationship to diminished cell adherence. The Journal of Clinical Investigation, 74, 536–551.CrossRefPubMedGoogle Scholar
  24. 24.
    Kaufman, D., Kilpatrick, L., Hudson, R. G., et al. (1999). Decreased superoxide production, degranulation, tumor necrosis factor alpha secretion, and CD11b/CD18 receptor expression by adherent monocytes from preterm infants. Clinical and Diagnostic Laboratory Immunology, 6, 525–529.PubMedGoogle Scholar
  25. 25.
    Jiang, Q., Azuma, E., Hirayama, M., et al. (2001). Functional immaturity of cord blood monocytes as detected by impaired response to hepatocyte growth factor. Pediatrics International, 43, 334–339.CrossRefPubMedGoogle Scholar
  26. 26.
    Pranke, P., Failace, R. R., Allebrandt, W. F., Steibel, G., Schmidt, F., & Nardi, N. B. (2001). Hematologic and immunophenotypic characterization of human umbilical cord blood. Acta Haematologica, 105, 71–76.CrossRefPubMedGoogle Scholar
  27. 27.
    Rainsford, E., & Reen, D. J. (2002). Interleukin 10, produced in abundance by human newborn t cells, may be the regulator of increased tolerance associated with cord blood stem cell transplantation. British Journal Haematology, 116, 702–709.CrossRefGoogle Scholar
  28. 28.
    Juretic, E., Gagro, A., Vukelic, V., & Petrovecki, M. (2004). Maternal and neonatal lymphocyte subpopulations at delivery and 3 days postpartum: Increased coexpression of cd45 isoforms. American Journal of Reproductive Immunology, 52, 1–7.CrossRefPubMedGoogle Scholar
  29. 29.
    De Paoli, P., Battistin, S., & Santini, G. F. (1988). Age-related changes in human lymphocyte subsets: progressive reduction of the CD4 CD45r (suppressor inducer) population. Clinical Immunology and Immunopathology, 48, 290–296.CrossRefPubMedGoogle Scholar
  30. 30.
    Xia, D., Hao, S., & Xiang, J. (2006). CD8+ cytotoxic T-APC stimulate central memory CD8+ T cell responses via acquired peptide-mhc class I complexes and CD80 costimulation, and IL-2 secretion. Journal of Immunology, 177, 2976–2984.Google Scholar
  31. 31.
    Hombach, A., Kohler, H., Rappl, G., & Abken, H. (2006). Human CD4+ T cells lyse target cells via granzyme/perforin upon circumvention of mhc class ii restriction by an antibody-like immunoreceptor. Journal of Immunology, 177, 5668–5675.Google Scholar
  32. 32.
    Diamond, A. S., & Gill, R. G. (2000). An essential contribution by IFN-gamma to CD8+ T cell-mediated rejection of pancreatic islet allografts. Journal of Immunology, 165, 247–255.Google Scholar
  33. 33.
    Cocchi, F., DeVico, A. L., Garzino-Demo, A., Arya, S. K., Gallo, R. C., & Lusso, P. (1995). Identification of RANTES, MIP-1 alpha, and MIP-1 beta as the major HIV- suppressive factors produced by CD8+ T cells. Science, 270, 1811–1815.CrossRefPubMedGoogle Scholar
  34. 34.
    Kim, J. J., Nottingham, L. K., & Sin, J. I. (1998). CD8 positive T cells influence antigen-specific immune responses through the expression of chemokines. The Journal of Clinical Investigation, 102, 1112–1124.CrossRefPubMedGoogle Scholar
  35. 35.
    Zigova, T., Willing, A. E., Saporta, S., et al. (2001). Apoptosis in cultured hNT neurons. Developmental Brain Research, 127, 63–70.CrossRefPubMedGoogle Scholar
  36. 36.
    Chen, Y., & Swanson, R. A. (2003). Astrocytes and brain injury. Journal of Cerebral Blood Flow and Metabolism, 23, 137–149.PubMedGoogle Scholar
  37. 37.
    Gao, Q., Li, Y., & Chopp, M. (2005). Bone marrow stromal cells increase astrocyte survival via upregulation of phosphinoside 3-kinase/threonine protein kinase and mitogen-activated protein kinase kinase/extracellular signal-regulated kinase pathways and stimulate astrocyte trophic factor gene expression after anaerobic insult. Neuroscience, 136, 123–134.CrossRefPubMedGoogle Scholar
  38. 38.
    Benavides, A., Pastor, D., Santos, P., Tranque, P. A., & Calvo, S. (2005). CHOP plays a pivotal role in the astrocyte death induced by oxygen and glucose deprivation. Glia, 52, 261–275.CrossRefPubMedGoogle Scholar
  39. 39.
    Schobitz, B., De Kloet, E. R., & Holsboer, F. (1994). Gene expression and function of interleukin 1, interleukin 6 and tumor necrosis factor in the brain. Progress in Neurobiology, 44, 397–432.CrossRefPubMedGoogle Scholar
  40. 40.
    Colasanti, M., Ramacci, M. T., Foresta, P., & Lauro, G. M. (1991). Different in vitro response to rIL-1 beta of newborn and adult rat astroglia. International Journal of Developmental Neuroscience, 9, 501–507.CrossRefPubMedGoogle Scholar
  41. 41.
    Goswami, S., Gupta, A., & Sharma, S. K. (1998). Interleukin-6-mediated autocrine growth promotion in human glioblastoma multiforme cell line U87MG. Journal of Neurochemistry, 71, 1837–1845.CrossRefPubMedGoogle Scholar
  42. 42.
    Hama, T., Kushima, Y., Miyamoto, M., Kubota, M., Takei, N., & Hatanaka, H. (1991). Interleukin-6 improves the survival of mesencephalic catecholaminergic and septal cholinergic neurons from postnatal, two-week-old rats in cultures. Neuroscience, 40, 445–452.CrossRefPubMedGoogle Scholar
  43. 43.
    Yamada, M., & Hatanaka, H. (1994). Interleukin-6 protects cultured rat hippocampal neurons against glutamate-induced cell death. Brain Research, 643, 173–180.CrossRefPubMedGoogle Scholar
  44. 44.
    Campbell, I. L., Abraham, C. R., Masliah, E., et al. (1993). Neurologic disease induced in transgenic mice by cerebral overexpression of interleukin 6. Proceedings of the National Academy of Sciences of the United States of America, 90, 10061–10065.CrossRefPubMedGoogle Scholar
  45. 45.
    Heyser, C. J., Masliah, E., Samimi, A., Campbell, I. L., & Gold, L. H. (1997). Progressive decline in avoidance learning paralleled by inflammatory neurodegeneration in transgenic mice expressing interleukin 6 in the brain. Proceedings of the National Academy of Sciences of the United States of America, 94, 1500–1505.CrossRefPubMedGoogle Scholar
  46. 46.
    Penkowa, M., Molinero, A., Carrasco, J., & Hidalgo, J. (2001). Interleukin-6 deficiency reduces the brain inflammatory response and increases oxidative stress and neurodegeneration after kainic acid-induced seizures. Neuroscience Letters, 102, 805–818.Google Scholar
  47. 47.
    Kotake, Y., Sang, H., Tabatabaie, T., Wallis, G. L., Moore, D. R., & Stewart, C. A. (2002). Interleukin-10 overexpression mediates phenyl-n-tert-butyl nitrone protection from endotoxemia. Shock, 17, 210–216.CrossRefPubMedGoogle Scholar
  48. 48.
    Dokka, S., Shi, X., Leonard, S., Wang, L., Castranova, V., & Rojanasakul, Y. (2001). Interleukin-10-mediated inhibition of free radical generation in macrophages. American Journal of Physiology. Lung Cellular and Molecular Physiology, 280, L1196–L1202.PubMedGoogle Scholar
  49. 49.
    Spera, P. A., Ellison, J. A., & Feuerstein, G. Z. (1998). IL-10 reduces rat brain injury following focal stroke. Neuroscience Letters, 251, 189–192.CrossRefPubMedGoogle Scholar
  50. 50.
    Fuchs, A. C., Granowitz, E. V., Shapiro, L., et al. (1996). Clinical, hematologic and immunologic effects of interleukin-10 in humans. Journal of Clinical Immunology, 16, 291–303.CrossRefPubMedGoogle Scholar
  51. 51.
    Davies, C. A., Loddick, S. A., Toulmond, S., Stroemer, R. P., Hunt, J., & Rothwell, N. J. (1999). The progression and topographic distribution of interleukin-1beta expression after permanent middle cerebral artery occlusion in the rat. Journal of Cerebral Blood Flow and Metabolism, 19, 87–98.PubMedGoogle Scholar
  52. 52.
    Lee, S. C., Liu, W., Dickson, D. W., Brosnan, C. F., & Berman, J. W. (1993). Cytokine production by human fetal microglia and astrocytes. Differential induction by lipopolysaccharide and IL-1 beta. Journal of Immunology, 150, 2659–2667.Google Scholar
  53. 53.
    Tomozawa, Y., Inoue, T., & Satoh, M. (1995). Expression of type I interleukin-1 receptor mRNA and its regulation in cultured astrocytes. Neuroscience Letters, 195, 57–60.CrossRefPubMedGoogle Scholar
  54. 54.
    Liu, M., Hurn, P. D., Roselli, C. E., & Alkayed, N. J. (2007). Role of p450 aromatase in sex-specific astrocytic cell death. Journal of Cerebral Blood Flow and Metabolism, 27, 135–141.CrossRefPubMedGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2010

Authors and Affiliations

  • Lixian Jiang
    • 1
    • 2
    • 3
  • Samuel Saporta
    • 1
    • 2
    • 3
  • Ning Chen
    • 1
    • 2
  • Cyndy Davis Sanberg
    • 5
  • Paul Sanberg
    • 1
    • 2
    • 3
  • Alison Willing
    • 1
    • 2
    • 3
    • 4
    Email author
  1. 1.Center for Excellence in Aging and Brain RepairUniversity of South FloridaTampaUSA
  2. 2.Department of Neurosurgery and Brain RepairUniversity of South FloridaTampaUSA
  3. 3.Department of Pathology and Cell BiologyUniversity of South FloridaTampaUSA
  4. 4.Department of Molecular Pharmacology & PhysiologyUniversity of South FloridaTampaUSA
  5. 5.Saneron CCEL Therapeutics, Inc.TampaUSA

Personalised recommendations