Morphological and functional differentiation in BE(2)-M17 human neuroblastoma cells by treatment with Trans-retinoic acid

Devon Andres,Betty Benton,Patrick M Mcnutt,Radharaman Ray,Kyle S Hubbard,John Petrali,Brian M Keyser

doi:10.1186/1471-2202-14-49

Abstract

BackgroundImmortalized neuronal cell lines can be induced to differentiate into more mature neurons by adding specific compounds or growth factors to the culture medium. This property makes neuronal cell lines attractive as in vitro cell models to study neuronal functions and neurotoxicity. The clonal human neuroblastoma BE(2)-M17 cell line is known to differentiate into a more prominent neuronal cell type by treatment with trans-retinoic acid. However, there is a lack of information on the morphological and functional aspects of these differentiated cells.ResultsWe studied the effects of trans-retinoic acid treatment on (a) some differentiation marker proteins, (b) types of voltage-gated calcium (Ca2+) channels and (c) Ca2+-dependent neurotransmitter ([3H] glycine) release in cultured BE(2)-M17 cells. Cells treated with 10 μM trans-retinoic acid (RA) for 72 hrs exhibited marked changes in morphology to include neurite extensions; presence of P/Q, N and T-type voltage-gated Ca2+ channels; and expression of neuron specific enolase (NSE), synaptosomal-associated protein 25 (SNAP-25), nicotinic acetylcholine receptor α7 (nAChR-α7) and other neuronal markers. Moreover, retinoic acid treated cells had a significant increase in evoked Ca2+-dependent neurotransmitter release capacity. In toxicity studies of the toxic gas, phosgene (CG), that differentiation of M17 cells with RA was required to see the changes in intracellular free Ca2+ concentrations following exposure to CG.ConclusionTaken together, retinoic acid treated cells had improved morphological features as well as neuronal characteristics and functions; thus, these retinoic acid differentiated BE(2)-M17 cells may serve as a better neuronal model to study neurobiology and/or neurotoxicity.

Highlights

Immortalized neuronal cell lines can be induced to differentiate into more mature neurons by adding specific compounds or growth factors to the culture medium
Morphological changes and possible synaptic activity To establish the conditions for retinoic acid (RA) treatment to induce cellular differentiation, we examined the effects of different RA concentrations (1, 5, and 10 μM) and times (48 – 120 hours)
BE(2)-M17 cell line (M17) cells transformed from an immature to a more mature neuronal state after 72 hours of exposure to 10 μM RA

Summary

Introduction

Immortalized neuronal cell lines can be induced to differentiate into more mature neurons by adding specific compounds or growth factors to the culture medium. This property makes neuronal cell lines attractive as in vitro cell models to study neuronal functions and neurotoxicity. There is a lack of information on the morphological and functional aspects of these differentiated cells Neurotoxic chemicals, such as lead (Pb) and organophosphorus (OP) insecticides are prevalent in the environment. The two main types of cell culture systems used for in vitro neurological testing are (a) primary neuronal cell cultures dissociated from peripheral or central nervous system tissues and (b) clonal cell lines derived from tumors of neurological origin [2]

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