Within each group, changes in MR data over time were examined by using ordinary least squares linear regression analyses

Sep 10, 2021 Other Apoptosis

Within each group, changes in MR data over time were examined by using ordinary least squares linear regression analyses. in vivoClabeled MSC transplants and unlabeled control transplants were E.coli polyclonal to V5 Tag.Posi Tag is a 45 kDa recombinant protein expressed in E.coli. It contains five different Tags as shown in the figure. It is bacterial lysate supplied in reducing SDS-PAGE loading buffer. It is intended for use as a positive control in western blot experiments compared by using tests. MR data were correlated with histopathologic results. Results: In vivoClabeled MSCs demonstrated significantly higher ferumoxytol uptake compared with ex vivoClabeled cells. With electron microscopy, iron oxide nanoparticles were localized in secondary lysosomes. In vivoClabeled cells demonstrated significant T2 shortening effects in BIBW2992 (Afatinib) vitro and in vivo when they were compared with unlabeled control cells (T2 in vivo, 15.4 vs 24.4 msec; BIBW2992 (Afatinib) < .05) and could be tracked in osteochondral defects for 4 weeks. Histologic examination confirmed the presence of iron in labeled transplants and defect remodeling. Conclusion: Intravenous ferumoxytol can be used to effectively label MSCs in vivo and can be used for tracking of stem cell transplants with MR imaging. This method eliminates risks of contamination and biologic alteration of MSCs associated with ex vivoClabeling procedures. ? RSNA, 2013 Supplemental material: (1C3). The need for knee replacement is rapidly increasing, with 3.48 million expected procedures by 2030 (4). However, artificial implants are associated with potential complications, such as periprosthetic fractures, loosening, and metal sensitivity (4C6). Even in the absence of complications, the lifetime of an artificial prosthesis is limited to approximately 10 years because of wear of the implant (7C9). Cell transplants, particularly stem cellCscaffold nanocomposites, overcome these problems by providing long-term biologic restoration of joint defects (10C14). Bone marrowCderived mesenchymal stem cells (MSCs) have been established as a promising source for stem cellCmediated joint repair in a clinical setting. MSCs can be easily obtained with a bone marrow aspirate, are efficiently expanded in vitro, and can differentiate into all joint components (15C17). However, interactions between transplanted MSCs and the patients host environment are still poorly understood. To monitor successful engraftment and recognize complications such as graft failure or tumor formation, MSC therapies require in vivo tracking of the transplanted stem cells with noninvasive imaging technologies. In the past, stem cell tracking has been achieved on the basis of the concept of ex vivo contrast agent labeling (18C23). This approach requires multiple ex vivo manipulations of stem cells between their harvest and transplantation. Clinical translation of ex vivoClabeling procedures is complicated from a regulatory point of view, as these manipulations greatly enhance the risk of cell sample contamination (24), alterations in stem cell biology, or in vivo side effects from added transfection agents (25C27). Most transfection agents (Lipofectamine 2000 [Invitrogen, Carlsbad, Calif] or poly-l-lysine [Sigma-P4707; Sigma-Aldrich, St Louis, Mo]) are not U.S. Food and Drug Administration (FDA) approved (28). In addition, some ultrasmall superparamagnetic iron oxideCtransfection agent combinations have induced cytotoxic effects (29C32) or altered the stem cell biology (33). To avoid these complications, we undertook to determine whether an immediately clinically applicable approach for stem cell labeling, which would not require ex vivo manipulations of harvested cells and which would eliminate the need for transfection agents, could be used to track transplanted MSCs. Our approach relies on intravenous administration of the FDA-approved iron supplement ferumoxytol (Feraheme; Advanced BIBW2992 (Afatinib) Magnetics, Cambridge, Mass) to a stem cell donor prior to stem cell harvest from bone marrow. Ferumoxytol is composed of iron oxide nanoparticles (34), which are taken up by the reticuloendothelial system in vivo (13,35C39) and which provide a strong signal intensity effect on magnetic resonance (MR) images (13,40C42). On the basis of these properties, we postulated that intravenously injected ferumoxytol would be taken up by MSCs in bone marrow, would be retained in the cells through harvesting and ex vivo expansion, and allow for sensitive in vivo MSC detection with MR imaging after transplantation into osteochondral defects. Thus, our aim was to determine whether intravenous ferumoxytol as a clinically applicable iron supplement can be used to effectively label MSCs in vivo and can be used for tracking of stem cell transplants. Materials and Methods In Vivo MSC Labeling The study was approved by the animal care and use committee at Stanford University (Stanford, Calif). Sixteen 6C8-week-old Sprague-Dawley rats (Charles River, Wilmington, Mass) served as MSC donors: Seven rats remained untreated, while nine rats were injected intravenously with ferumoxytol (= 7) or fluorescein isothiocyanate (FITC) (Fisher Scientific, Pittsburgh,.