Orthogonal potency analysis of mesenchymal stromal cell function during ex vivo expansion

Abstract

Adult bone marrow mesenchymal stromal cells (MSCs) have cross-functional, intrinsic potency that is of therapeutic interest. Their ability to regenerate bone, fat, and cartilage, modulate the immune system, and nurture the growth and function of other bone marrow hematopoietic stem/progenitor cells have all been evaluated by transplant applications of MSCs. These applications require the isolation and expansion scaled cell production. To investigate biophysical properties of MSCs that can be feasibly utilized as predictors of bioactivity during biomanufacturing, we used a low-density seeding model to drive MSCs into proliferative stress and exhibit the hallmark characteristics of in vitro aging. A low-density seeding method was used to generate MSCs from passages 1-7 to simulate serial expansion of these cells to maximize yield from a single donor. MSCs were subjected to three bioactivity assays in parallel to ascertain whether patterns in MSC age, size, and shape were associated with the outcomes of the potency assays. MSC age was found to be a predictor of adipogenesis, while cell and nuclear shape was strongly associated to hematopoietic-supportive potency. Together, these data evaluate morphological changes associated with cell potency and highlight new strategies for purification or alternatives to assessing MSC quality.

Molecular Mechanisms Contributing to Mesenchymal Stromal Cell Aging

“…An in vitro model of low-density MSC growth was for example recently used to evaluate the effect of age on a series of MSC biophysical properties used as predictors of bioactivity. The findings of the study indicated that MSC age is a predictor of adipogenesis, while cell and nuclear shape are strongly associated to hematopoietic-supportive potency [204].…”

Cryopreservation Impacts Cell Functionality of Long Term Expanded Adipose-Derived Stem Cells

“…There is convincing evidence that the duration of culture expansion influenced ADSC phenotype and ascertained the relationship between morphological changes and physiological effect [22,[40][41][42] which might explain the mechanisms whereby ADSC ensure tissue repair. Nevertheless, cryopreservation effect on expanded ADSC physiology should not be excluded adding to the complexity of their physiologymicroenvironment interactions.…”

Exosome: A New Player in Translational Nanomedicine

“…Despite the usefulness of these criteria for identifying MSCs, these cell populations are reported to be heterogenous with regard to their non-clonal proliferation, differentiation potencies, stromal stem cell profiles and committed progenitors [117,118]. These differences appear to be related to their tissue origin and to the cell separation and culture expansion techniques used [119,120]. Thus, the complex composition of the exosomes released is markedly influenced by initial local cell crosstalk and microenvironmental priming.…”