Cryopreservation of embryonic stem cell‐derived multicellular neural aggregates labeled with micron‐sized particles of iron oxide for magnetic resonance imaging

Abstract

Magnetic resonance imaging (MRI) provides an effective approach to track labeled pluripotent stem cell (PSC)-derived neural progenitor cells (NPCs) for neurological disorder treatments after cell labeling with a contrast agent, such as an iron oxide derivative. Cryopreservation of pre-labeled neural cells, especially in three-dimensional (3D) structure, can provide a uniform cell population and preserve the stem cell niche for the subsequent applications. In this study, the effects of cryopreservation on PSC-derived multicellular NPC aggregates labeled with micron-sized particles of iron oxide (MPIO) were investigated. These NPC aggregates were labeled prior to cryopreservation because labeling thawed cells can be limited by inefficient intracellular uptake, variations in labeling efficiency, and increased culture time before use, minimizing their translation to clinical settings. The results indicated that intracellular MPIO incorporation was retained after cryopreservation (70-80% labeling efficiency), and MPIO labeling had little adverse effects on cell recovery, proliferation, cytotoxicity and neural lineage commitment post-cryopreservation. MRI analysis showed comparable detectability for the MPIO-labeled cells before and after cryopreservation indicated by T2 and T2* relaxation rates. Cryopreserving MPIO-labeled 3D multicellular NPC aggregates can be applied in in vivo cell tracking studies and lead to more rapid translation from preservation to clinical implementation.

Functionalization of Brain Region-specific Spheroids with Isogenic Microglia-like Cells

“…Approximately 2 × 10 5 microglia-like cells (day 30), hiPSC-derived neural progenitor cells (iNPCs) (day 21) and hiPSC-derived endothelial cells (iECs) (day 21) 76 were replated onto 24-well plate in DMEM plus 10% FBS. The adherent cells were incubated with 1 ml fresh DMEM plus 10% FBS containing 0.25 × 10 8 fluorescent (0.86 μm, flash red, 660/690 nm) micron-sized particles of iron oxide (MPIO)/mL (Bangs Laboratories, Fishers, IN, USA, Part number ME03F/9772) corresponding to the concentration at 2.5 μg Fe/mL 77,78 . The attached microglia-like cells that were not labeled with MPIO served as control.…”

Pluripotent stem cell expansion and neural differentiation in 3-D scaffolds of tunable Poisson’s ratio

“…Murine ES-D3 line (American Type Culture Collection, Manassas, VA) was maintained on 0.1% gelatin-coated 6-well plates in a standard 5% CO 2 incubator as previously reported [29].…”

Cryopreservation of Induced Pluripotent Stem Cell-Derived Dopaminergic Neurospheres for Clinical Application

“…Several groups have reported the effects of cryopreservation on neurospheres ( Supplementary Table 3 ). They all found good cell viability in vitro , but the in vivo survival and function in the brain are unknown [ 33–36 ]. In most cases, the spheres were cryopreserved at – (0.5–1)°C/min using DMSO as a CPA based on previous reports for the cryopreservation of neural tissues [ 37 ].…”