Bone marrow cell transplantation time-dependently reverses G-CSF effects after stroke in hypertensive rats

Granulocyte colony-stimulating factor (G-CSF) is a hematopoietic cytokine and preclinically proven neuroprotectant. A potential reason for the clinical failure of G-CSF may be that relevant G-CSF effects such as the mobilization of mononuclear hematopoietic stem/progenitor cells from the bone marrow may take too long in humans (up to 9 days) to counter initial stroke consequences. Systemic transplantation of bone marrow mononuclear cells (BMMNCs) is feasible within a relatively short time after stroke onset and may provide an external resource of aforementioned stem/progenitor cells, thereby ""bridging the gap"" until G-CSF comes to full effect. Male spontaneously hypertensive rats (SHR) were randomly assigned into four groups after permanent middle cerebral artery occlusion (MCAO). Groups 1-3 received IP G-CSF treatment (50 mg/kg) for 5 days starting 1 h after stroke onset. Groups 2 and 3 also received 1.5 × 107/kg BMMNCs IV at 6 or 48 h following stroke, respectively. Group 4 received placebo treatment. Functional deficits (adhesive removal test), infarct volume, and edema (T2 TSE MRI) were repeatedly assessed for 1 month. Peripheral leukocyte counts and BMMNC biodistribution were analyzed by flow cytometry during the first week after stroke. G-CSF monotreatment reduced functional deficits (p < 0.05) and partially reversed poststroke immune depression [overall leuko-/monocyte as well as B-, natural killer (NK), and T-cell counts; p < 0.01] and as expected increased peripheral leukocyte counts massively (p < 0.01). G-CSF did not affect infarct volume or edema. BMMNC cotransplantation at 6 h did not further improve functional deficits (p > 0.05 each). Surprisingly, BMMNC transplantation at 48 h abolished G-CSF effects. Early biodistribution studies (at 52 h after stroke onset) revealed splenic accumulation of granulocytes and BMMNCs as well as a granulocyte overload in the peripheral circulation and the brain (p < 0.05). Splenic accumulation of transplanted BMMNCs may have impaired peripheral granulocyte clearance. Subsequently, increased granulocyte numbers in the circulation and the poststroke brain prompted a proinflammatory bias of the innate immune system's response to stroke, ultimately abolishing G-CSF effects. These surprising findings indicate that systemic effects of experimental stroke therapies need to be carefully considered when assessing the therapeutic potential of such novel approaches.