Homing of Stem Cells in Stroke Therapeutics

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DESCRIPTION (provided by applicant): Somatic stem cells appear to have their greatest impact on stroke therapeutics not by literally replacing cells but by protecting host neural cells & circuitry from progressive damage as well as catalyzing endogenous host regenerative responses. When such stem cell-mediated actions have been invoked, behavioral improvements have been seen. Neuroectoderm-derived stem cells ("neural stem cells [NSCs]") have been the "gold-standard" for such therapeutics against which any alternative stem cell type must be compared. However, a number of labs have found efficacy from the intravascular administration of human umbilical cord blood-derived stem cells (hUCSCs). The intravascular administration of stem cells allows one to circumvent more invasive neurosurgical implantation, assuming that the cells can efficiently home to the region of injury. An additional advantage of cell-mediated protective therapies (as opposed to pharmacological-mediated interventions) is that the window of opportunity can be as wide as 24-72 hrs, making them well-suited to the realities of when patients with stroke present to health care facilities. Although UCSCs have been promising, the ability to extend their use to actual patients has practical & immunological limitations. In particular, based on extrapolating from animal studies, adult patients would require combining multiple cord blood units in order to obtain sufficient numbers of stem cells to achieve efficacy. Furthermore, combining units places stress on identifying compatible units with no more than 1-2 HLA mismatches to minimize an immune reaction. Also, efficient delivery of cells to where they are most needed & not to areas where they are not required or could even create problems - i.e., homing - is pivotal to the success of all cell-based therapeutics for any pathological condition. UCSCs must have optimal access to injured host cells & their milieu if efficacy & safety are to be maximized. When considering systemic administration of either UCSCs or NSCs for mediating brain repair following stroke, rolling & adhesion on endothelial cells is the critical 1st step in the homing cascade that is necessary for targeting these cells to the injured brain. Most hUCSCs exhibit deficient rolling & adhesion on endothelial cells, severely limiting their therapeutic potential. NSCs have a similar limitation. Recent studies have shown that this deficit can be corrected by pretreating hUCSCs with fucosyltransferase-VI which fucosylates sLeX & completes the P-selectin glycoprotein ligand (PSGL) on the CD34+ cells. The net effect is to increase recognition of the PSGL for selectins on endothelial cells. It is unknown whether this works, however, for brain vascular endothelium. It is also unknown whether this works for NSCs -- again, the "gold standard" -- although they, too, have an incomplete unfucosylated selectin ligand which compromises their homing. We propose first [Aim 1] to determine in vitro the optimal parameters for fucosylating & thereby enhancing the binding of hUCSCs & human NSCs (hNSCs) to inflamed human brain-derived endothelial cells under physiological shear stress (employing a specialized flow chamber). Subsequently [Aim 2], we will use these conditions to administer fucosylated (vs. control) hUCSCs & hNSCs to a rat model of stroke, comparing the 2 stem cell types (for the 1st time) head-to-head. Rats will be injected iv with the stem cells at 24-72 hrs post-middle cerebral artery occlusion. Outcome measures will include behavior; infarct volume (by MRI); spectroscopy (by MRS); migration (by MRI & bioluminescence imaging); sparing of host neurons & their connections (by immunohistochemistry & tract tracing). These data will be correlated with histology. These studies could help provide a strategy for enhancing the homing of stem cells to the brain in a minimally-invasive manner, using cell numbers & a time frame that is practical for providing clinical benefit in acute/subacute stroke, particularly if the stem cells serve to protect extant neural tissue & connections; diminish inflammation, scarring & secondary injury processes; detoxify the milieu; & promote neovascularization. Such studies could also provide insights that extend to other types of CNS pathology & other types of stem cells. PUBLIC HEALTH RELEVANCE: Peripheral intravascular administration of either human umbilical cord-derived stem cells (hUCSCs) or neural stem cells (hNSCs) for acute stroke therapeutics in adults requires a sufficient number of cells with fully expressed homing mechanisms for optimal targeting to the area of compromise. This proposal will assess a new technology that could expand the use of available supplies of either source of stem cells. The process of placing a sugar group - a fucosyl group - on stem cell membranes through a simple enzymatic pre-treatment process has shown in preclinical studies to increase homing of hUCSCs and likely will extend to hNSCs as well. Aim 1 will identify the optimal conditions for preparing the stem cells while Aim 2 will comparatively assess in a rat stroke model the efficacy of hUCSCs vs. hNSCs following peripheral administration 24 hrs post-stroke, particularly under conditions where their homing has been optimized by fucosylation.