The Effect of Coatings and Nerve Growth Factor on Attachment and Differentiation of Pheochromocytoma Cells.

Anna Orlowska,Vladimir Baulin,Russell Crawford,Elena Ivanova,Shahram Ghanaati,Mohammad Al Kobaisi,Huu Nguyen,Pallale Perera,Andre Dias

doi:10.3390/ma11010060

Abstract

Cellular attachment plays a vital role in the differentiation of pheochromocytoma (PC12) cells. PC12 cells are noradrenergic clonal cells isolated from the adrenal medulla of Rattus norvegicus and studied extensively as they have the ability to differentiate into sympathetic neuron-like cells. The effect of several experimental parameters including (i) the concentration of nerve growth factor (NGF); (ii) substratum coatings, such as poly-L-lysine (PLL), fibronectin (Fn), and laminin (Lam); and (iii) double coatings composed of PLL/Lam and PLL/Fn on the differentiation process of PC12 cells were studied. Cell morphology was visualised using brightfield phase contrast microscopy, cellular metabolism and proliferation were quantified using a 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium (MTS) assay, and the neurite outgrowth and axonal generation of the PC12 cells were evaluated using wide field fluorescence microscopy. It was found that double coatings of PLL/Lam and PLL/Fn supported robust adhesion and a two-fold enhanced neurite outgrowth of PC12 cells when treated with 100 ng/mL of NGF while exhibiting stable metabolic activity, leading to the accelerated generation of axons.

Highlights

The pheochromocytoma (PC12) cell line is commonly used in in vitro studies to examine neuronal differentiation and neurotoxicity implicated in neurodegenerative disease [1,2]
Single- and two-component coatings were investigated in this study
Since PLL was reported to facilitate cell attachment and improve the differentiation of PC12 cells [24], PLL was selected for analysis

Summary

Introduction

The pheochromocytoma (PC12) cell line is commonly used in in vitro studies to examine neuronal differentiation and neurotoxicity implicated in neurodegenerative disease [1,2]. PC12 cells are noradrenergic clonal cells originating from Rattus norvegicus transplantable pheochromocytoma [1]. They exhibit a reversible response to nerve growth factor (NGF). After NGF exposure, PC12 cells acquire characteristic phenotypic properties associated with sympathetic neuron-like cells, which includes the inhibition of proliferation, outgrowth of neurites, and the possibility of being electrically excitable [1,2,3]. The neuron-like PC12 cells start to express various integral proteins that are responsible for neurite growth [1] and can transmit signals along the axons [4,5].

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