Microcarrier expansion of c-MycERTAM -modified human olfactory mucosa cells for neural regeneration.

Ana Valinhas,Ivan B Wall,Gerardo Santiago‐Toledo

doi:10.1002/bit.27573

Abstract

Human olfactory mucosa cells (hOMCs) have potential as a regenerative therapy for spinal cord injury. In our earlier work, we derived PA5 cells, a polyclonal population that retains functional attributes of primary human OMCs. Microcarrier suspension culture is an alternative to planar two-dimensinal culture to produce cells in quantities that can meet the needs of clinical development. This study aimed to screen the effects of 10 microcarriers on PA5 hOMCs yield and phenotype. Studies performed in well plates led to a 2.9-fold higher cell yield on plastic compared to plastic plus microcarriers with upregulation of neural markers β-III tubulin and nestin for both conditions. Microcarrier suspension culture resulted in concentrations of 1.4 × 105 cells/ml and 4.9 × 104 cells/ml for plastic and plastic plus, respectively, after 7 days. p75NTR transcript was significantly upregulated for PA5 hOMCs grown on Plastic Plus compared to Plastic. Furthermore, coculture of PA5 hOMCs grown on Plastic Plus with a neuronal cell line (NG108-15) led to increased neurite outgrowth. This study shows successful expansion of PA5 cells using suspension culture on microcarriers, and it reveals competing effects of microcarriers on cell expansion versus functional attributes, showing that designing scalable bioprocesses should not only be driven by cell yields.

Highlights

This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record
And recent studies have attributed the neural regeneration of the Human olfactory mucosa cells (hOMCs) population to the presence of several different cell types, such as neural stem cells (NSCs), mesenchymal stem cells (MSCs), olfactory neurons and olfactory ensheathing cells (OECs), a type of glia
We report the microcarrier expansion of a candidate cell population for the treatment of spinal cord injury (SCI), PA5 hOMCs, on microcarrier stirred culture using spinner flasks, with the aim to identify a microcarrier type that maximize the expansion of neuroprotective cell types such as OECs, NSCs, and MSCs, which can lead to increased potency

Summary

Introduction

This article has been accepted for publication and undergone full peer review but has not been through the copyediting, typesetting, pagination and proofreading process, which may lead to differences between this version and the Version of Record. The generation of a conditionallyimmortalized hOMC population, such as PA5 hOMCs, enables a potentially extended life span, allowing the application of a cell-banking model-based manufacturing process, necessary for an allogeneic cell therapy The translation of such a therapeutic product to the market would require the development of a scalable bioprocess, able to yield large amounts of cells which can reach doses up to 1x107 cells/dose (Casarosa et al, 2014). We report the microcarrier expansion of a candidate cell population for the treatment of SCI, PA5 hOMCs, on microcarrier stirred culture using spinner flasks, with the aim to identify a microcarrier type that maximize the expansion of neuroprotective cell types such as OECs, NSCs, and MSCs, which can lead to increased potency. We report the use of a microcarrier-based spinner flasks system for the expansion of the PA5 hOMCs population of cells, a candidate cell line for the treatment of spinal cord-injury. The qPCR reactions were performed in Bio-Rad Hard-Shell Low Profile Thin-Wall 96 Well Skirted PCR plates on the Bio-Rad

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