Nicolas Bon-Mardion

Comparative gene expression profiling of olfactory ensheathing cells from olfactory bulb and olfactory mucosa.

Olfactory ensheathing cells (OEC) have the ability to promote regeneration in the nervous system. Hence, they hold promise for cell therapy. Most of the experimental studies have investigated the role of OECs taken from olfactory bulb (OB). However, for a clinical human application, olfactory mucosa (OM) seems to be the only acceptable source for OECs. Many studies have compared the distinct ability of OECs from OB and OM to improve functional nerve regeneration after lesion of the nervous system. Nevertheless, the two populations of OECs may differ in several points, which might affect all fate after transplantation in vivo. We report here the first study which compares gene expression profiling between these two populations of OECs. It appears that OB‐OECs and OM‐OECs display distinct gene expression pattern, which suggest that they may be implicated in different physiological processes. Notably, OM‐OECs overexpress genes characteristic of wound healing and regulation of extra cellular matrix. In contrast, OB‐OECs gene profile suggests a prominent role in nervous system development. Hence, OB‐OECs and OM‐OECs fundamentally differ in their gene expression pattern, which may represent a crucial point for future clinical application.

Co-transplantation of olfactory ensheathing cells from mucosa and bulb origin enhances functional recovery after peripheral nerve lesion.

Olfactory ensheathing cells (OECs) represent an interesting candidate for cell therapy and could be obtained from olfactory mucosa (OM-OECs) or olfactory bulbs (OB-OECs). Recent reports suggest that, depending on their origin, OECs display different functional properties. We show here the complementary and additive effects of co-transplanting OM-OECs and OB-OECs after lesion of a peripheral nerve. For this, a selective motor denervation of the laryngeal muscles was performed by a section/anastomosis of the recurrent laryngeal nerve (RLN). Two months after surgery, recovery of the laryngeal movements and synkinesis phenonema were analyzed by videolaryngoscopy. To complete these assessments, measure of latency and potential duration were determined by electrophysiological recordings and myelinated nerve fiber profiles were defined based on toluidine blue staining. To explain some of the mechanisms involved, tracking of GFP positive OECs was performed. It appears that transplantation of OM-OECs or OB-OECs displayed opposite abilities to improve functional recovery. Indeed, OM-OECs increased recuperation of laryngeal muscles activities without appropriate functional recovery. In contrast, OB-OECs induced some functional recovery by enhancing axonal regrowth. Importantly, co-transplantation of OM-OECs and OB-OECs supported a major functional recovery, with reduction of synkinesis phenomena. This study is the first which clearly demonstrates the complementary and additive properties of OECs obtained from olfactory mucosa and olfactory bulb to improve functional recovery after transplantation in a nerve lesion model.

Transplantation of olfactory ensheathing cells promotes axonal regeneration and functional recovery of peripheral nerve lesion in rats

Introduction: Olfactory ensheathing cells (OECs) hold promise for cell therapy because they may promote regeneration of the central nervous system. However, OECs have been less studied after peripheral nerve injury (PNI). The purpose of this investigation was to determine the effect of OEC transplantation on a severe sciatic nerve (SN) lesion. Methods: OECs were injected in rats after section and 2‐cm resection of the SN. Results: Three months after therapy, muscle strength and morphometric studies showed complete restoration of the contractile properties of the gastrocnemius and complete repair of the SN. Immunohistochemistry and RT‐PCR studies indicated an increase in the presence of neurotrophic factors. Interestingly, tracking of green fluorescent protein (GFP)‐positive OECs showed that no OECs were present in the SN. Discussion: Our results demonstrate that, after severe PNI, OECs have remarkable potential for nerve regeneration by creating a favorable microenvironment. Muscle Nerve, 2011

Potential of Olfactory Ensheathing Cells from Different Sources for Spinal Cord Repair

Spinal cord injury (SCI) induces a permanent disability in patients. To this day no curative treatment can be proposed to restore lost functions. Therefore, extensive experimental studies have been conducted to induce recovery after SCI. One of the most promising therapies is based on the use of olfactory ensheathing cells (OECs). OECs can be obtained from either the olfactory bulbs (OB-OECs) or from olfactory mucosa (OM-OECs), involving a less invasive approach for autotransplantation. However the vast majority of experimental transplantations have been focusing on OB-OECs although the OM represents a more accessible source of OECs. Importantly, the ability of OM-OECs in comparison to OB-OECs to induce spinal cord recovery in the same lesion paradigm has never been described. We here present data using a multiparametric approach, based on electrophysiological, behavioral, histological and magnetic resonance imaging experiments on the repair potential of OB-OECs and OM-OECs from either primary or purified cultures after a severe model of SCI. Our data demonstrate that transplantation of OECs obtained from OB or OM induces electrophysiological and functional recovery, reduces astrocyte reactivity and glial scar formation and improves axonal regrowth. We also show that the purification step is essential for OM-OECs while not required for OB-OECs. Altogether, our study strongly indicates that transplantation of OECs from OM represents the best benefit/risk ratio according to the safety of access of OM and the results induced by transplantations of OM-OECs. Indeed, purified OM-OECs in addition to induce recovery can integrate and survive up to 60 days into the spinal cord. Therefore, our results provide strong support for these cells as a viable therapy for SCI.

Efficiency of laryngeal motor nerve repair is greater with bulbar than with mucosal olfactory ensheathing cells.

The real ability of OECs provided by olfactory mucosa cultures (OM-OECs) and those from olfactory bulb cultures (OB-OECs) must be better characterized in order to propose their future clinical application. Therefore, we used a lesion of the vagus nerve (VN), which constitutes a severe motor denervation due to long distance of the muscular targets (4.5 cm). We performed a section/anastomosis surgery of the VN, at the third tracheal ring. Then, OM-OECs and OB-OECs were injected in matrigel around the lesion site. Three months after surgery, laryngeal muscle activity, synkinesis phenomena and latency were evaluated by videolaryngoscopy and electromyography recordings. To complete these procedures, axonal morphometric study of the right recurrent nerve was performed to assess axonal regrowth and tracking of green fluorescent protein positive cells was performed. Recurrent nerve is the motor branch innervating the laryngeal muscles, and is located distally to the lesion, near the muscular targets (0.7 cm). These analyses permitted to compare the ability of these two populations to improve functional recovery and axonal regrowth. Our results show that, OM-OECs improved electrical muscular activity and nervous conduction with significant tissue healing but induced aberrant movement and poor functional recovery. In contrast, OB-OECs induced a partial functional recovery associated with an increase in the number of myelinated fibers and nervous conduction. Our study suggests that, as recently reported in a microarray study, OM-OECs and OB-OECs express different properties. In particular, OM-OECs could regulate inflammation processes and extracellular matrix formation but have a poor regeneration potential, whereas, OB-OECs could improve functional recovery by inducing targeted axonal regrowth.

Involvement of the aero-digestive tract in swallowing-ventilation coordination: an animal study

Background  The precise coordination of swallowing and ventilation is considered an important mechanism for minimizing the risk of pulmonary aspiration. The aim of this study was to test the hypothesis that the aero‐digestive tract is involved in swallowing‐ventilation coordination in rats.

Transplantation of olfactory ensheathing cells to evaluate functional recovery after peripheral nerve injury.

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth and regrowth of the primary olfactory neurons. Indeed, the primary olfactory system is characterized by its ability to give rise to new neurons even in adult animals. This particular ability is partly due to the presence of OECs which create a favorable microenvironment for neurogenesis. This property of OECs has been used for cellular transplantation such as in spinal cord injury models. Although the peripheral nervous system has a greater capacity to regenerate after nerve injury than the central nervous system, complete sections induce misrouting during axonal regrowth in particular after facial of laryngeal nerve transection. Specifically, full sectioning of the recurrent laryngeal nerve (RLN) induces aberrant axonal regrowth resulting in synkinesis of the vocal cords. In this specific model, we showed that OECs transplantation efficiently increases axonal regrowth. OECs are constituted of several subpopulations present in both the olfactory mucosa (OM-OECs) and the olfactory bulbs (OB-OECs). We present here a model of cellular transplantation based on the use of these different subpopulations of OECs in a RLN injury model. Using this paradigm, primary cultures of OB-OECs and OM-OECs were transplanted in Matrigel after section and anastomosis of the RLN. Two months after surgery, we evaluated transplanted animals by complementary analyses based on videolaryngoscopy, electromyography (EMG), and histological studies. First, videolaryngoscopy allowed us to evaluate laryngeal functions, in particular muscular cocontractions phenomena. Then, EMG analyses demonstrated richness and synchronization of muscular activities. Finally, histological studies based on toluidine blue staining allowed the quantification of the number and profile of myelinated fibers. All together, we describe here how to isolate, culture, identify and transplant OECs from OM and OB after RLN section-anastomosis and how to evaluate and analyze the efficiency of these transplanted cells on axonal regrowth and laryngeal functions.

Effects of chronic aspirations on breathing pattern and ventilatory drive in vagatomized rats.

The aim of this study was to test the hypothesis that aspirations induced by unilateral vagotomy destabilise ventilatory pattern during swallowing. The study was carried out on 15 Wistar rats (2-3 months, 290-350 g) using whole-body plethysmography and video recordings, before and after unilateral vagotomy. The rats were given water ad libitum via a baby bottle fitted with a nipple. The experiment was continued until rest ventilation and swallowing periods were identified on the video recordings. Following the sectioning of the right vagus nerve, all the rats presented bronchial aspirations and unilateral vocal cord paralysis in the aperture position. After the vagotomy there were no changes at rest of the ventilatory variables compared to healthy controls. In healthy animals during swallowing, we observed a decrease in total ventilatory time (TTOT), a decrease in inspiratory time (TI) (p < 0.001), a decrease in expiratory time (TE) (p < 0.001), no change in tidal volume (VT) and an increase in mean inspiratory time (VT/TI) (p < 0.001) compared to the rest period. Animals with chronic aspiration presented during swallowing an increase in TTOT (p < 0.001), TI (p < 0.01), and TE (p < 0.001), no change in VT and a decrease of VT/TI (p < 0.001) and a modification of ventilatory pattern. In conclusion, our results confirmed that swallowing modifies ventilation in healthy animals and that chronic aspiration decreases ventilatory drive and modifies ventilatory pattern during swallowing.

Dual innervation may occur in a partially denervated muscle: Dual innervation of a muscle

Introduction: With a view to simplifying surgical techniques for selective laryngeal reinnervation, we addressed the question of whether it is feasible to receive additional innervation by a partially denervated muscle using an infrahyoid muscle model. Methods: In 90 rats (6 groups of 15), phrenic nerve transfer was used to reinnervate the sternothyroid muscle. In some cases, residual innervation by the original nerve was present. Three months later we performed electromyographic studies, contraction strength measurements, histologic assessment, and retrograde labeling. Results: Muscles reinnervated by the phrenic nerve had a greater “dual‐response” rate (in terms of nerve latency, contraction strength, and retrograde labeling) than muscles in the control groups. Discussion: The phrenic nerve can impart its inspiratory properties to an initially denervated strap muscle—even when residual innervation is present. The preservation of contractile potential confirmed the feasibility of dual innervation in a previously injured muscle. Muscle Nerve 59:108–115, 2019