Single-cell high-content imaging parameters predict functional phenotype of cultured human bone marrow stromal stem cells

Abstract

Abstract Cultured human bone marrow stromal (mesenchymal) stem cells (hBM-MSCs) are heterogenous cell populations exhibiting variable biological properties. Quantitative high-content imaging technology allows identification of morphological markers at a single cell resolution that are determinant for cellular functions. We determined the morphological characteristics of cultured primary hBM-MSCs and examined their predictive value for hBM-MSC functionality. BM-MSCs were isolated from 56 donors and characterized for their proliferative and differentiation potential. We correlated these data with cellular and nuclear morphological features determined by Operetta; a high-content imaging system. Cell area, cell geometry, and nucleus geometry of cultured hBM-MSCs exhibited significant correlation with expression of hBM-MSC membrane markers: ALP, CD146, and CD271. Proliferation capacity correlated negatively with cell and nucleus area and positively with cytoskeleton texture features. In addition, in vitro differentiation to osteoblasts as well as in vivo heterotopic bone formation was associated with decreased ratio of nucleus width to length. Multivariable analysis applying a stability selection procedure identified nuclear geometry and texture as predictors for hBM-MSCs differentiation potential to osteoblasts or adipocytes. Our data demonstrate that by employing a limited number of cell morphological characteristics, it is possible to predict the functional phenotype of cultured hBM-MSCs and thus can be used as a screening test for “quality” of hBM-MSCs prior their use in clinical protocols. Significance statement Clinical trials employing cultured human bone marrow mesenchymal stem cells (hBM-MSCs) demonstrate the presence of a large inter-donor variability in biological functions and clinical efficacy. By applying high-content imaging methodology to cultured BM-MSCs obtained from a large cohort of donors, morphological features including cell area and nucleus geometry as predictors for cell differentiation and proliferation were identified. These parameters can serve as selection criteria for BM-MSC populations to be used in clinical trials of enhancing bone regeneration.