Detection and protection of the neurovascular unit (NVU) are essential for treatment of acute stroke patients, especially the use of tissue plasminogen activator (tPA). In the present study, we conducted in vivo and ex vivo optical imaging for detecting activation of matrix metalloproteinases (MMPs) and evaluated the protective effect of intracerebral transplantation of bone marrow stromal cells (BMSCs) obtained from green fluorescent protein (GFP) transgenic (Tg) mice on the NVU in tPA-mediated brain damage after transient middle cerebral artery occlusion (tMCAO) in mice. Compared with the tMCAO group, the tMCAO plus BMSC group showed significant reductions of in vivo and ex vivo fluorescent signals for MMPs at 48 hr after tMCAO, with a partial colocalization of BMSC-GFP signals. Intracerebrally transplanted BMSCs ameliorated MMP-9 activation by immunohistochemistry and Western blot with differentiation into microglial and astroglial cells. Double-immunofluorescence study revealed improved NVU disruption in the tMCAO plus BMSC group. The present study suggests that intracerebral BMSC transplantation reduced MMP activation and subsequent NVU disruption caused by tPA after tMCAO and that this MMP activation and BMSC effect were detectable with in vivo and ex vivo optical imaging.