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Stem cell therapy modulates not only the local microenvironment of the brain but also the systemic immune responses. We explored the impact of human multipotent adult progenitor cells (MAPC) modulating splenic activation and peripheral immune responses after ischemic stroke. Hundred twenty-six Long-Evans adult male rats underwent middle cerebral artery occlusion. Twenty-four hours later, they received IV MAPC or saline treatment. At 3 days after infusion, RNA was isolated from the injured cortex and spleen for microarray analysis. Spleen mass, splenocyte phenotype, and releasing cytokines were measured. Serum cytokines, MAPC biodistribution, brain lesion sizes and neurofunctional deficits were compared in rats treated with MAPC or saline with and without spleens. Stroked animals treated with MAPC exhibited genes that more closely resembled animals with sham surgery. Gene categories downregulated by MAPC included leukocyte activation, antigen presentation, and immune effector processing, associated with the signaling pathways regulated by TNF-α, IL-1β, IL-6, and IFN-γ within the brain. MAPC treatment restored spleen mass reduction caused by stroke, elevated Treg cells within the spleen, increased IL-10 and decreased IL-1β released by splenocytes. MAPC reduced IL-6 and IL-1β and upregulated IL-10 serum levels. Compared with saline, MAPC enhance stroke recovery in rats with intact spleens but had no effects in rats without spleens. MAPC restores expression of multiple genes and pathways involved in immune and inflammatory responses after stroke. Immunomodulation of the splenic response by the intravenous administration of MAPC may create a more favorable environment for brain repair after stroke. Stem Cells 2017;35:1290-1302.

Although mononuclear cells (MNCs) from bone marrow are being investigated in phase I clinical trials in stroke patients, dose response, therapeutic time window and biodistribiton have not been well-characterized in animal stroke models. Long Evans rats underwent common carotid artery/middle cerebral artery occlusion (CCA/MCAo) and 24 hrs later were randomized to receive saline IV or a bone marrow aspiration followed by an IV infusion of autologous separated MNCs (1 million, 10 million or 30 million cells/kg). In another experiment, rats underwent CCAo/MCAo and were randomized at 24 hrs, 72 hrs or 7 days after stroke to receive a saline injection or 10 million/kg MNCs. All animals were evaluated on the cylinder and corner tests up to 28 days. MNCs were tracked using Q-dot nanocrystals to monitor biodistribution. Animals treated with MNCs at 10 million and 30 million cells/kg at 24 hrs after stroke had significant reductions in neurological deficits and lesion size compared to saline controls or animals treated with 1 million cells/kg. There was no difference in neurological deficits in the 10 and 30 million cell/kg groups at 28 days. Animals treated with MNCs at 72 hrs but not at 7 days showed a significant reduction in neurological deficits by 28 days. Labeled MNCs were found in the brain, spleen, lung, liver, and kidney at 1 hr and exponentially decreased over the ensuing week. In conclusion, we found a maximum reduction in neurological deficits at 10 and 30 million cells/kg and a therapeutic time window up to 72 hrs after stroke.

Background and Purpose Bone marrow derived mononuclear cells (MNCs) are an investigational autologous cell-based therapy for acute ischemic stroke. Both intravenous (IV) and intra-arterial (IA) administration routes have been used in clinical trials. However, the route of administration to optimize the effect of MNCs is unknown. In this study, we compared the effect of IV versus IA route of MNCs in the rat stroke model. Methods Long Evans rats were subjected to transient middle cerebral artery occlusion (MCAo). At 24 hrs after stroke, animals were randomly assigned to either receive autologous bone marrow derived MNCs using IV or IA delivery routes. IV saline served as control. 1 million cells/kg (low dose) and 30 million cells/kg (high dose) were assessed. Neurological testing, cavity size, serum cytokines, neuroregenerative endpoints, and MNC biodistribution were evaluated. Results High dose MNCs improved functional recovery, reduced lesion size and pro-inflammatory cytokines, and increased vessel density and neurogenesis markers compared with saline treatment (p<0.05). However, there were no significant differences between IV and IA MNC treated groups; though, IV MNCs reduced serum IL-1β levels compared with IA MNCs (p<0.05). IA MNCs at high dose led to a greater number of cells in the brain at 1 and 6 hrs after injection but not in the lungs and spleen. Low dose MNCs (by IV or IA) did not improve any functional or structural endpoint compared with saline. Conclusion At low and high doses of MNCs, we found that IV or IA achieves similar structural and functional outcomes after stroke.

Autologous bone marrow-derived mononuclear cells (MNCs) are a potential therapy for ischemic stroke. However, the effect of MNCs in intracerebral hemorrhage (ICH) has not been fully studied. In this study, we investigated the effects of autologous MNCs in experimental ICH. ICH was induced by infusion of autologous blood into the left striatum in young and aged male Long Evans rats. Twenty-four hours after ICH, rats were randomized to receive an intravenous administration of autologous MNCs (1 · 10 7 cells/kg) or saline. We examined brain water content, various markers related to the integrity of the neurovascular unit and inflammation, neurological deficit, neuroregeneration, and brain atrophy. We found that MNC-treated young rats showed a reduction in the neurotrophil infiltration, the number of inducible nitric oxide synthase-positive cells, and the expression of inflammatory-related signalings such as the high-mobility group protein box-1, S100 calcium binding protein B, matrix metalloproteinase-9, and aquaporin 4. Ultimately, MNCs reduced brain edema in the perihematomal area compared with saline-treated animals at 3 days after ICH. Moreover, MNCs increased vessel density and migration of doublecortin-positive cells, improved motor functional recovery, spatial learning, and memory impairment, and reduced brain atrophy compared with saline-treated animals at 28 days after ICH. We also found that MNCs reduced brain edema and brain atrophy and improved spatial learning and memory in aged rats after ICH. We conclude that autologous MNCs can be safely harvested and intravenously reinfused in rodent ICH and may improve long-term structural and functional recovery after ICH. The results of this study may be applicable when considering future clinical trials testing MNCs for ICH.