Nitric Oxide Facilitates Delivery and Mediates Improved Outcome of Autologous Bone Marrow Mononuclear Cells in a Rodent Stroke Model

Bone marrow mononuclear cells (MNC) represent an investigational treatment for stroke. The objective of this study was to determine the relevance of vasoactive mediators, generated in response to MNC injection, as factors regulating cerebral perfusion (CP), the biodistribution of MNC, and outcome in stroke.Long Evans rats underwent transient middle cerebral artery occlusion. MNC were extracted from the bone marrow at 22 hrs and injected via the internal carotid artery or the femoral vein 2 hours later. CP was measured with MRI or continuous laser Doppler flowmetry. Serum samples were collected to measure vasoactive mediators. Animals were treated with the Nitric Oxide (NO) inhibitor, L-NAME, to establish the relevance of NO-signaling to the effect of MNC. Lesion size, MNC biodistribution, and neurological deficits were assessed.CP transiently increased in the peri-infarct region within 30 min after injecting MNC compared to saline or fibroblast control. This CP increase corresponded temporarily to serum NO elevation and was abolished by L-NAME. Pre-treatment with L-NAME reduced brain penetration of MNC and prevented MNC from reducing infarct lesion size and neurological deficits.NO generation in response to MNC may represent a mechanism underlying how MNC enter the brain, reduce lesion size, and improve outcome in ischemic stroke

Single shot T1rho magnetic resonance imaging of metabolically generated water in vivo

The use of Oxygen-17 MRI provides great promise for the clinically-useful quantification of metabolism. To bring techniques based on 17O closer to clinical application, we demonstrate imaging of metabolically generated H2 17O in pigs after 17O2 delivery with increased temporal resolution T1rho-weighted imaging and precision delivery of 17O2 gas. The kinetics of the appearance of H2 17O in pig brains are displayed with one to two minutes of 17O2 delivery, the shortest delivery times reported in the literature. It is also shown that H2 17O concentrations can be quantified with single shot T1rho imaging based on a balanced steady state free precession readout, and that with this strategy pausing to reduce T1 saturation increases sensitivity to H2 17O over acquisition in the steady state. Several additional considerations with this sequence, which can be generalized to any pre-encoding cluster, such as energy deposition are considered

Details of different behavioral tests and scores.

<p>Details of different behavioral tests and scores.</p

Illustration of a pulmonary artery of an animal pre-treated with L-NAME or Saline before IV MNC administration.

<p>Arrows point toward Q tracker labeled MNCs.</p

Biodistribution of MNC in brain, spleen and lungs after IV administration of MNCs(10 million) at 24 hrs after stroke.

<p>MNCs were labeled with Q-tracker(green). These animals had undergone stroke, 24 hrs later were injected with MNCs IV and then sacrificed 1 hr afterwards. Sections were prepared at 1 hr after injection and counterstained with DAPI (blue). L-NAME was injected via IP 1 hr prior to MNC administration. N = 3 per group. (A): Representative microscopic pictures illustrating MNCs (green+blue) in animals pre-treated with L-NAME (top row) or saline (bottom row). Magnification: 400×. (B): A histogram illustrating the mean number of labeled MNCs per high power field in the brain, spleen, and lungs. Animals were pre-treated with L-NAME before IV injection of MNCs or animals were pre-treated with saline followed by IV injection of MNCs (‡p<0.05).</p

Time course of changes of NO at 24 hours after stroke.

<p>(A): Serum NO levels after saline or MNC (IV or IA) administered at 24 hrs post stroke. §p<0.05 for IA or IV MNC treated groups compared with the saline treated group. Time zero refers to the time right before cell or saline injection. N = 5 per group. (B): Serum NO levels in animals treated with L-NAME administered 60 min before MNC injection IV. §p<0.05 refers to the MNC group compared with the saline group; ‡ p<0.05 refers to the MNC group compared with the L-NAME+MNC group. N = 5 per group.</p

Experimental design.

<p>Experimental design.</p

Changes in Cerebral perfusion.

<p>(A) Perfusion weighted imaging was performed in animals receiving MNC or saline at 24 hrs after MCA occlusion. There was a significant decrease (‡p<0.05) in the mean TTP in the peri-infarct region at 5 minutes after MNC infusion via IA or IV routes of delivery. There was no significant change in TTP in the saline group before and after injection. N = 4 per group. (B) Cerebral perfusion (CP) was monitored with Laser Doppler flowmetry over the peri-infarct area of animals administered MNCs (10 million cells) or saline at 24 hrs after MCA occlusion. CP was expressed after injection as a percentage compared with the baseline pre-injection value. CP increased at 5 minutes after MNC injection. There was no significant change in CP in the saline treated group. N = 6 per group. (C): Time course of changes in cerebral perfusion after IV injection of MNCs (10 million cells), FBCs (10 million), or saline. ‡p<0.05 refers to the MNC group compared to the saline group and §p<0.05 refers to the MNC group compared with the FBC group. There was significant elevation of CP after MNC infusion at 1, 3, 5, 10 and 15 minutes. There was no significant change in CP in the saline or FBC groups. ‘0’ on x-axis represents the time in minutes, just before the injection. N = 6 per group.</p