A2B5-positive oligodendrocyte precursor cell transplantation improves neurological deficits in rats following spinal cord contusion associated with changes in expression of factors involved in the Notch signaling pathway

Abstract

Background

Oligodendrocyte precursor cells (OPCs) are myelinated glial cells of the central nervous system (CNS), able to regenerate oligodendrocytes and myelin. This study aimed to elucidate the effect of A2B5-positive (A2B5+) OPC transplantation in rats with spinal cord contusion (SCC) and to investigate changes in expression of various factors involved in the Notch signaling pathway after OPC transplantation.

Methods

OPCs were obtained from induced pluripotent stem cells (iPSCs) originating from mouse embryo fibroblasts (MEFs). After identification of iPSCs and iPSC-derived OPCs, A2B5+ OPCs were transplanted into the injured site of rats with SCC one week after SCC insult. Behavioral tests evaluated motor and sensory function 7 days after OPC transplantation. Real-time quantitative polymerase chain reaction (RT-qPCR) determined the expression of various cytokines related to the Notch signaling pathway after OPC transplantation.

Results

IPSC-derived OPCs were successfully generated from MEFs, as indicated by positive immunostaining of A2B5, PDGFα and NG2. Further differentiation of OPCs was identified by immunostaining of Olig2, Sox10, Nkx2.2, O4, MBP and GFAP. Importantly, myelin formation was significantly enhanced in the SCC+ OPC group and SCI-induced motor and sensory dysfunction was largely alleviated by A2B5+ OPC transplantation. Expression of factors involved in the Notch signaling pathway (Notch-1, Numb, SHARP1 and NEDD4) was significantly increased after OPC transplantation.

Conclusions

A2B5+ OPC transplantation attenuates motor and sensory dysfunction in SCC rats by promoting myelin formation, which may be associated with change in expression of factors involved in the Notch signaling pathway.

Introduction

Spinal cord injury (SCI) is a serious disease resulting from partial or completely loss of neuronal functions [1]. The social and psychological problems in patients caused by SCI requires costly medical care [2]. SCI, caused by the culmination of tissue damage, is characterized by multitudinous neurological dysfunctions [3]. The primary damage of cells at the injury site is followed by a cascade of secondary processes which include inflammation, oxidative stress, ischemia of surrounding tissues, increased blood barrier permeability, demyelination, calcium overload, glial scar formation, excitotoxicity, and axonal degeneration [4], [5]. Currently, the clinical treatment of spinal cord injury (SCI) includes surgical intervention, pharmacological agents and cellular therapies. However, the neuroprotective effects of nonsurgical treatments remain obscure and need more investigations in SCI.

Stem cell therapies have been utilized in SCI animal models for decades and applied in human bodies in recent years, which has shown some progress of recovery [6]. Different types of cells have been tested, including mesenchymal stromal cells, glial cells, pluripotent stem cells derived neural cells and neural stem cells [7]. Embryonic stem cells (ESCs) have been employed for the treatment of SCI, and many transplanted ESCs show differentiation into oligodendrocytes [8]. Another study showed ESCs can be pre-differentiated into oligodendrocyte lineage [9], and the transplanted human ESCs-derived oligodendrocyte precursor cells promote SCI myelin reformation and restoration of motor function [10]. IPSCs can be obtained by artificially inducing non-pluripotent cells to express certain genes. IPSCs display a lot of similarities in many aspects with embryonic stem cells (ESCs), such as the expression of gene and protein, teratoma, formation, doubling, embryo formation, chromatin methylation patterns, different chimera formation and differentiation potentials [11]. Vanessa M et al. proposed that the application of iPSCs could demonstrate a wonderful interface in disease modeling, regenerative medicine and developmental biology [12]. IPSCs technology enables people to escape from the ethical debate and therapeutic cloning fierce in solution, also avoids the problem of immune cell transplantation rejection. IPSCs technology provides new prospects for basic research on the application of cell transplantation in the treatment of human diseases. Oligodendrocyte precursor cells (OPCs) is a kind of cell surface ganglioside epitope expressing in developing thymic epithelial cells, neuroendocrine cells and oligodendrocyte progenitors [13], [14]. Alireza Pouya et al. [15] found a cell population of oligodendrocyte progenitors from human induced pluripotent stem cells (hiPSCs) by embryoid body formation. It has been reported that OPCs derived from hiPSCs were transplanted into the thrombolytic induced optic chiasma of rats, and the dysfunctions were largely alleviated [16]. However, few have been reported about the potential of OPCs for SCI therapy.

Notch signaling pathway, a hetero-oligomer composed of extracellular portion, is calcium-dependent and has non-covalent interaction with a smaller fraction of the notch protein consisting of a extracellular region, a single transmembrane-pass, and a small intrastitial region [17]. Notch signaling pathway is a conserved cell signaling system in organisms, affecting many processes of normal cell morphogenesis, including differentiation of pluripotent progenitor cells, cell apoptosis, cell proliferation and cell boundary formation [18]. The Notch signaling pathway also induces long-term effects on the neighboring cells, including neuroglia and neurons during nervous development [19]. The proliferative signaling could be promoted by Notch signaling during neurogenesis, and the activities of Notch signaling are inhibited by Numb for promoting neural differentiation. The Notch signaling pathway is known as an effective pathway regulating the recovery of vascular formation and capillary functional morphology. Its role in SCI remains to be elucidated. Based on the mentioned scientific problems, this study aimed to determine the effect of the OPCs transplantation in SCC rats and investigate the expression variation of various factors in Notch signaling pathway, providing preliminary mechanistic basis for the OPCs transplantation alleviating SCI.