Marrow Stromal Cell Infusion Rescues Hematopoiesis in Lethally Irradiated Mice despite Rapid Clearance after Infusion

Abstract

Marrow stromal cells (MSCs, also termed mesenchymal stem cells) have been proposed as a promising cellular therapy for tissue injury including radiation-induced marrow failure, but evidence for a direct effect is lacking. To assess the effects of MSCs on survival after lethal irradiation, we infused syngeneic MSCs (either as immortalized MSCs clones or primary MSCs) intravenously into wild-type C57/Bl6 mice within 24 hours of lethal total body irradiation (TBI). Mice receiving either of the MSC preparations had significantly improved survival when compared to controls. In vivo imaging, immune histochemistry, and RT-PCR employed to detect MSCs indicated that the infused MSCs were predominantly localized to the lungs and rapidly cleared following infusion. Our results suggest that a single infusion of MSCs can improve survival after otherwise lethal TBI but the effect is not due to a direct interaction with, or contribution to, the damaged marrow by MSCs.

Figure 1

Impact of MSC infusion on survival of C57/Bl6 mice. (a) Determination of lethal dose of radiation for C57/Bl6 mice when radiation delivered with the RS2000 X-ray irradiator. Adult female mice 6 to 8 weeks in age were radiated with 600, 700, or 800 cGy radiation (n = 5 each) in two divided doses survival rate calculated at 7 weeks after radiation. None of the animals that received 700 or 800 cGy radiation survived past 4 weeks from day of radiation. 4 of the 5 mice in the 600 cGy radiation group survived past 7 weeks from day of radiation (P < 0.001). (b) Survival of mice receiving MSCs (clones or primary) when compared to control animals. Survival was studied in adult female mice receiving 700 cGy radiation followed by one of the following interventions: cloned MSCs (cMSC, n = 19), primary MSCs (pMSC, n = 20) or PBS (n = 21). Survival was significantly better for the cMSC (53%) and pMSC (60%) groups when compared to PBS (P values of 0.017 and 0.041, resp. with logrank test) at 7 weeks after radiation.

Figure 2

Tissue distribution of MSCs in vivo after injection. A. Bioluminescent imaging (BLI) of MSCs at 4, 7, and 24 hours after injection. 1 × 10⁶ MSC cells stably expressing ffluc were infused into recipient mice by tail vein injection followed by Bioluminescent signals were restricted to the thoracic area. Signals decreased rapidly and were undetectable 72 hours after infusion. B. Quantitative RT-PCR for ffluc in the above tissues harvested at 1, 4, and 7 days after infusion (n = 6 each). Whole RNA was prepared from tissues and analyzed for presence of ffluc mRNA by q RT-PCR. Since ffluc expression cannot be normalized to any endogenous control, equal quantities of starting material (2 μg of whole RNA) were used for cDNA synthesis. Those samples with mean Ct < 35 cycles were deemed as positive for ffluc expression. All animals had detectable levels of ffluc mRNA in their lungs on day 1, while most other tissues had no detectable transcript on day 1. On days 4 and 7, there were detectable levels of ffluc transcripts detectable in a variable proportion of all tissues from some animals.