Homing of annexin-labeled stem cells to apoptotic cells

Abstract

Ischemic diseases are characterized by the presence of pro-apoptotic stimuli, which initiate a cascade of processes that lead to cell injury and death. Several molecules and events represent detectable indicators of the different stages of apoptosis. Among these indicators is phosphatidylserine (PS) translocation from the inner to the outer leaflet of the plasma membrane, which can be detected by annexinV (ANXA5) conjugation. This is a widely used in vivo and in vitro assay marking the early stages of apoptosis. We report here on an original method that employs PS-ANXA5 conjugation to target stem cells to apoptotic cells. Mesenchymal stem cells (MSCs) from GFP-positive transgenic rats were biotinylated on membrane surfaces with sulfosuccinimidyl-6-(biotinamido) hexanoate (sulfo-NHS-LC-biot) and then bound to avidin. The avidin-biotinylated MSCs were labeled with biotin conjugated ANXA5. Bovine aortic endothelial cells (BAE-1 cells) were exposed to UVC to induce caspasedependent apoptosis. Finally, we tested the ability of ANXA5-labeled MSCs to Unauthenticated Download Date | 5/11/18 2:02 AM CELLULAR & MOLECULAR BIOLOGY LETTERS 101 bind BAE-1 apoptotic cells: suspended ANXA5-labeled MSCs were seeded for 1 hour on a monolayer of UV-treated or control BAE-1 cells. After washing, the number of MSCs bound to BAE-1 cells was evaluated by confocal microscopy. Statistical analysis demonstrated a significant increase in the number of MSCs tagged to apoptotic BAE-1 cells. Therefore, stem cell ANXA5 tagging via biotin-avidin bridges could be a straightforward method of improving homing to apoptotic tissues.

Methods to assess stem cell lineage, fate and function

“…Thus far, these probes have been used mainly for cancer imaging, but these techniques can be potentially applied to detect apoptosis after stem cell delivery (63). Interestingly, a recent study demonstrated that conjugation of annexin to stem cells improved homing to apoptotic cells in vitro (64). Further investigation is needed to better define the utility of these techniques for stem cell therapy.…”