Mannitol enhances delivery of marrow stromal cells to the brain after experimental intracerebral hemorrhage

Abstract

Previous studies show that intravascular injection of human bone marrow stromal cells (hBMSCs) significantly improves neurological functional recovery in a rat model of intracerebral hemorrhage (ICH). In the present study, we tested the hypothesis that mannitol improves the efficiency of intraarterial MSC delivery (i.e., fewer injected cells required for therapeutic efficacy) after ICH. There were four post-ICH groups (N=9): group 1, negative control with only intraarterial injection of 1 million human fibroblasts in phosphate-buffered saline (PBS); group 2, intravenous injection of mannitol alone in PBS (1.5 g/kg); group 3, intraarterial injection of 1 million hBMSCs alone in PBS; and group 4, intravenous injection of mannitol (1.5 g/kg) in PBS followed by intraarterial injection of 1 million hBMSCs in PBS. Group 4 exhibited significantly improved neurological functional outcome as assessed by neurological severity score (NSS) and corner test scores. Immunohistochemical staining of group 4 suggested increased synaptogenesis, proliferating immature neurons, and neuronal migration. The number of hBMSCs recruited to the injured region increased strikingly in group 4. Tissue loss was notably reduced in group 4. In summary, the beneficial effects of intraarterial infusion of MSCs are amplified with intravenous injection of mannitol. Preadministration of mannitol significantly increases the number of hBMSCs located in the ICH region, improves histochemical parameters of neural regeneration, and reduces the anatomical and pathological consequences of ICH.

Fig. 1

Neurological functional tests and striatal tissue loss percentage of the injured side. Quantitative bar graph results of NSS (left panel) and corner turn test (middle panel) of four groups (control, FB; mannitol, MT; hBMSC; combination treatment, hBMSC+MT) are presented in the left and middle panels. Bar graphs of quantitative striatal tissue loss percentages in the ICH region relative to the contralateral normal region of four groups are shown in right panel. Statistical significance level is: *P<0.05.

Fig. 2

A representative H&E stained coronal section at the level of the anterior commissure of rat brain. The detected areas of the injured hemisphere recognized under the microscope are the hemorrhage core area (red), the boundary zone (yellow), and the subventricular zone (SVZ, pink).

Fig. 3

H&E staining showing representative sections of fibroblast control group (A), mannitol group (B), hBMSC group (C) and the combination group (D).

Fig. 4

Immunostaining of the control and combination treatment sections showing the quantitative immunoreactivities of mAb 1281, synaptophysin, and DCX. The other two groups are not shown. Note that a higher number of cells per area are stained to mAb 1281, synaptophysin and DCX. Quantitative immunoreactivities for all four groups are presented as bar graphs on the right side of each panel. Scale bar = 50 um.

Fig. 5

Immunostaining of the control and combination treatment sections showing the quantitative immunoreactivities of BrdU and TUJ1. Quantitative immunoreactivities for all four groups are presented as bar graphs on the right side of each panel. Colocalization of BrdU and TUJ1 in a subpopulation of cells near the injured region of the combination group is shown on the bottom panel. Arrows indicate cells positively stained for both BrdU and TUJ1. Scale bar = 50 um.