ARPE-19 conditioned medium promotes neural differentiation of adipose-derived mesenchymal stem cells

Abstract

BACKGROUND Adipose-derived stem cells (ASCs) have been increasingly explored for cell-based medicine because of their numerous advantages in terms of easy availability, high proliferation rate, multipotent differentiation ability and low immunogenicity. In this respect, they have been widely investigated in the last two decades to develop therapeutic strategies for a variety of human pathologies including eye disease. In ocular diseases involving the retina, various cell types may be affected, such as Müller cells, astrocytes, photoreceptors and retinal pigment epithelium (RPE), which plays a fundamental role in the homeostasis of retinal tissue, by secreting a variety of growth factors that support retinal cells. AIM To test ASC neural differentiation using conditioned medium (CM) from an RPE cell line (ARPE-19). METHODS ASCs were isolated from adipose tissue, harvested from the subcutaneous region of healthy donors undergoing liposuction procedures. Four ASC culture conditions were investigated: ASCs cultured in basal Dulbecco's Modified Eagle Medium (DMEM); ASCs cultured in serum-free DMEM; ASCs cultured in serum-free DMEM/F12; and ASCs cultured in a CM from ARPE-19, a spontaneously arising cell line with a normal karyotype derived from a human RPE. Cell proliferation rate and viability were assessed by crystal violet and MTT assays at 1, 4 and 8 d of culture. At the same time points, ASC neural differentiation was evaluated by immunocytochemistry and western blot analysis for typical neuronal and glial markers: Nestin, neuronal specific enolase (NSE), protein gene product (PGP) 9.5, and glial fibrillary acidic protein (GFAP). RESULTS Depending on the culture medium, ASC proliferation rate and viability showed some significant differences. Overall, less dense populations were observed in serum-free cultures, except for ASCs cultured in ARPE-19 serum-free CM. Moreover, a different cell morphology was seen in these cultures after 8 d of treatment, with more elongated cells, often showing cytoplasmic ramifications. Immunofluorescence results and western blot analysis were indicative of ASC neural differentiation. In fact, basal levels of neural markers detected under control conditions significantly increased when cells were cultured in ARPE-19 CM. Specifically, neural marker overexpression was more marked at 8 d. The most evident increase was observed for NSE and GFAP, a modest increase was observed for nestin, and less relevant changes were observed for PGP9.5. CONCLUSION The presence of growth factors produced by ARPE-19 cells in tissue culture induces ASCs to express neural differentiation markers typical of the neuronal and glial cells of the retina.

Machine Learning Elucidates Design Features of Plasmid DNA Lipid Nanoparticles for Cell Type-Preferential Transfection

“…The two exceptions were N2a and ARPE-19 cells, which were the most highly correlated cell lines with a Spearman's rank correlation coefficient of 0.71. We attributed the high correlation to the shared neuronal origins of ARPE-19 and N2a cells 31,32 . Our results suggest that it may be possible to exploit interactions between LNP design features and certain cellular features of the target cell type to develop tailored LNPs for cell type-preferential transfection through modulation of the formulation compositions and helper lipid identity.…”

Surface Functionalized Polyaniline Nanofibers:Chitosan Nanocomposite for Promoting Neuronal-like Differentiation of Primary Adipose Derived Mesenchymal Stem Cells and Urease Activity

“…In addition, adipose-derived MSCs were shown to have long-term stability under culture conditions with higher proliferative capacity. Lately, several reports have been focused on using ADMSCs for nerve regeneration/neurological disorders. − Combining all of the advantages discussed above, present study demonstrates a simple one step and cost-efficient surface functionalization approach of PNF:Ch nanocomposites using an optimized concentration of glutaraldehyde (GA) to elevate the adhesion and neuronal differentiation potential of primary ADMSCs under the differentiation protocol based on bFGF and EGF.…”

Serum-Free Cultures: Could They Be a Future Direction to Improve Neuronal Differentiation of Mesenchymal Stromal Cells?

“…Further studies are needed to clarify the mechanisms by which AD-MSCs differentiate towards a neural phenotype. In this regard, the presence of extracellular vesicles in ARPE-19 CM cannot be excluded [ 79 ].…”