Trophic factor induction of human umbilical cord blood cells in vitro and in vivo.

The mononuclear fraction of human umbilical cord blood (HUCBmnf) is a mixed cell population that multiple research groups have shown contains cells that can express neural proteins. In these studies, we have examined the ability of the HUCBmnf to express neural antigens after in vitro exposure to defined media supplemented with a cocktail of growth and neurotrophic factors. It is our hypothesis that by treating the HUCBmnf with these developmentally-relevant factors, we can expand the population, enhance the expression of neural antigens and increase cell survival upon transplantation. Prior to growth factor treatment in culture, expression of stem cell antigens is greater in the non-adherent HUCBmnf cells compared to the adherent cells (p < 0.05). Furthermore, treatment of the non-adherent cells with growth factors, increases BrdU incorporation, especially after 14 days in vitro (DIV). In HUCBmnf-embryonic mouse striata co-culture, a small number of growth factor treated HUCBmnf cells were able to integrate into the growing neural network and express immature (nestin and TuJ1) and mature (GFAP and MAP2) neural markers. Treated HUCBmnf cells implanted in the subventricular zone predominantly expressed GFAP although some grafted HUCBmnf cells were MAP2 positive. While short-term treatment of HUCBmnf cells with growth and neurotrophic factors enhanced proliferative capacity in vitro and survival of the cells in vivo, the treatment regimen employed was not enough to ensure long-term survival of HUCBmnf-derived neurons necessary for cell replacement therapies for neurodegenerative diseases.

Stroke-induced migration of human umbilical cord blood cells: time course and cytokines.

The therapeutic window for treatment of individuals after stroke is narrow, regardless of the treatment regime; extension of this window would provide a major therapeutic advance. In prior reports, we demonstrated significant improvements in the behavioral defects of rats that received human umbilical cord blood (HUCB) cells 24 h after a middle cerebral arterial occlusion. These effects paralleled the recruitment of these cells to the site of tissue damage. While the administration of HUCB cells 24 h after stroke was effective, the optimal time to administer these cells after stroke has not been established. Here, we investigated the migration of HUCB cells to ischemic tissue extracts. After ischemic assault, brain tissue was homogenized, and the supernatants were assayed for their ability to attract HUCB mononuclear cells as well as for levels of several cytokines. We demonstrate increased migratory activity of HUCB cells toward the extracts harvested at 24-72 h after stroke. The extracts possessed increased levels of certain cytokines and chemokines, suggesting their participation in HUCB cell migration. The results from this study are promising in that the current 3-h therapeutic window for the treatment of stroke victims, using approved anticoagulant treatment, may be extended with the use of HUCB cell therapy 24-72 h post stroke. Last, the chemokines present in the supernatant provide a sound starting point to start examining the mechanisms responsible for the in vivo migration of HUCB cells after the induction of stroke.

Transplantation of human umbilical cord blood cells benefits an animal model of Sanfilippo syndrome type B.

Sanfilippo syndrome type B is caused by alpha-N-acetylglucosaminidase (Naglu) enzyme deficiency leading to an accumulation of undegraded heparan sulfate, a glycosaminoglycan (GAG). Cell therapy is a promising new treatment and human umbilical cord blood (hUCB) cell transplantation may be preferred for delivery of the missing enzyme. We investigated the ability of mononuclear hUCB cells administered into the lateral cerebral ventricle to ameliorate/prevent histopathological changes in mice modeling Sanfilippo syndrome type B. These are the first results supporting enzyme replacement by administered hUCB cells. In vivo, transplanted hUCB cells survived long-term (7 months), migrated into the parenchyma of the brain and peripheral organs, expressed neural antigens, and exhibited neuron and astrocyte-like morphology. Transplant benefits were also demonstrated by stable cytoarchitecture in the hippocampus and cerebellum, and by reduced GAGs in the livers of treated mutant mice. A hUCB cell transplant may be an effective therapeutic strategy for enzyme delivery in Sanfilippo syndrome type B.

Human Umbilical Cord Blood Cells Have Trophic Effects on Young and Aging Hippocampal Neurons in Vitro.

In experimental models of central nervous system (CNS) aging, injury and disease, administering human umbilical cord blood (HUCB) cells induce recovery, most likely by interacting with multiple cellular processes. The aim of this study was to examine whether the HUCB cells produce trophic factors that may enhance survival and maturation of hippocampal neurons in an in vitro test system. We co-cultured the mononuclear fraction of HUCB cells with hippocampal neurons isolated from either young (7-months of age) or aging (21 month of age) rat brain for 14, 21, 28, 35 and 42 days in vitro (DIV), respectively. Immunocytochemistry was then employed to identify neurons (MAP2(+)) and glial cells (GFAP(+)) as well as arborization of neurites. The average number of MAP2(+) hippocampal neurons cells in both young and aging neuronal-HUCB co-cultures was significantly higher than in the control cultures (hippocampal mono-cultures). These MAP2(+) neurons in co-culture were richly arborized, especially in 21 and 28 DIV co-cultures, and expressed functional enzymes (Synaptophysin, tyrosine hydryoxlase (TH)), gamma amino butyric acid receptor (GABAAr) and glutamate transporter (EAAC1). The majority of hippocampal neurons in both co-culture systems grew very well and survived for up to 42 DIV with an increment of immature neurons which were positive for Nestin and TuJ1. Using a multiplex protein array, a number of secreted proteins that could have trophic effects on the neurons were identified.

Cord blood rescues stroke-induced changes in splenocyte phenotype and function.

The neuroprotective mechanism of human umbilical cord blood cells (HUCBC) in the rat middle cerebral artery occlusion (MCAO) stroke model remains uncertain. Given the inflammatory sequelae that occur following stroke, we investigated whether HUCBC protection could be derived from the modulation of this immuno-inflammatory event, suggested by the attraction of the HUCBC to the spleen. We found that, following MCAO, rat spleen size was reduced concomitantly with their CD8+ T-cell counts. Interestingly, MCAO-induced spleen size reduction correlated with the extent of ischemic damage, however, HUCBC treatment rescued the spleen weight, splenic CD8+ T-cell counts, as well as the amount of brain injury. Additionally, splenocyte proliferation assays demonstrated that HUCBC treatment opposed MCAO-associated T-cell proliferation by increasing the production of IL-10 while decreasing IFN-gamma. Taken together, these results suggest a novel immunomodulatory mechanism by which HUCBC mediate protection in the rat MCAO model of stroke.