Abstract
BackgroundPeripheral nerve injury is a worldwide clinical problem, and the preferred surgical method for treating it is the end-to-end neurorrhaphy. When it is not possible due to a large nerve gap, autologous nerve grafting is used. However, these surgical techniques result in nerve regeneration at highly variable degrees. It is thus very important to seek complementary techniques to improve motor and sensory recovery. One promising approach could be cell therapy. Transplantation therapy with human embryonic stem cells (hESCs) is appealing because these cells are pluripotent and can differentiate into specialized cell types and have self-renewal ability. Therefore, the main objective of this study was to find conditions under which functional recovery is improved after sciatic nerve neurorrhaphy. We assumed that hESC, either alone or in combination with heterologous fibrin sealant scaffold, could be used to support regeneration in a mouse model of sciatic nerve injury and repair via autografting with end-to-end neurorrhaphy.MethodsFive millimeters of the sciatic nerve of C57BL/6 J mice were transected off and rotated 180 degrees to simulate an injury, and then stumps were sutured. Next, we applied heterologous fibrin sealant and/or human embryonic stem cells genetically altered to overexpress fibroblast growth factor 2 (FGF2) at the site of the injury. The study was designed to include six experimental groups comprising neurorrhaphy (N), neurorrhaphy + heterologous fibrin sealant (N + F), neurorrhaphy + heterologous fibrin sealant + doxycycline (N + F + D), neurorrhaphy + heterologous fibrin sealant + wild-type hESC (N + F + W), neurorrhaphy + heterologous fibrin sealant + hESC off (N + F + T), and neurorrhaphy + heterologous fibrin sealant + hESC on via doxycycline (N + F + D + T). We evaluated the recovery rate using Catwalk and von Frey functional recovery tests, as well as immunohistochemistry analysis.ResultsThe experiments indicated that sensory function improved when transgenic hESCs were used. The regeneration of sensory fibers indeed led to increased reflexes, upon stimulation of the paw ipsilateral to the lesion, as seen by von-Frey evaluation, which was supported by immunohistochemistry.ConclusionsOverall, the present data demonstrated that transgenic embryonic stem cells, engineered to overexpress FGF-2 in an inducible fashion, could be employed to support regeneration aiming at the recovery of both motor and sensory functions.
Highlights
Peripheral nerve injury is a worldwide clinical problem, and the preferred surgical method for treating it is the end-to-end neurorrhaphy
We considered to include electronic pressure-meter test to our experiments
Expression of FGF-2 by human embryonic stem cells (hESC) We set a demonstration experiment in the lab to ensure that the addition of doxycycline activates the bioengineered cells to overexpress FGF-2
Summary
Peripheral nerve injury is a worldwide clinical problem, and the preferred surgical method for treating it is the end-to-end neurorrhaphy. If direct end-toend repair is not possible, due to lengthy nerve injury and retraction of the stumps, autografting using sensory donor nerves is the gold standard approach. When the gap is above the critical size, which is about 1 cm in rats, a graft is needed to bridge the damaged ends, reconnecting the proximal and distal stumps [4] In such situations, ‘autogenous nerve grafting’ is considered as the standard clinical treatment [1, 5]. In this grafting technique, a comparable nerve is first removed from another part of the patient’s body and is used to bridge the gap and connect the two ends of the severed nerve [6, 7]. These injuries will possibly never heal and can be permanently debilitating [4]
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