Combination of heterologous fibrin sealant and bioengineered human embryonic stem cells to improve regeneration following autogenous sciatic nerve grafting repair

 

Roghayeh Mozafari¹, Sergiy Kyrylenko², Mateus Vidigal Castro¹, Rui Seabra Ferreira Jr³, Benedito Barraviera³, Alexandre Leite Rodrigues Oliveira¹

 

1 Department of Structural and Functional Biology, Institute of Biology, University of Campinas (UNICAMP), Laboratory of Nerve Regeneration, Campinas, SP CEP 13083-970, Brazil.

2 Department of Public Health, Medical Institute of Sumy State University, Sumy 40007, Ukraine.

3 Center for the Study of Venoms and Venomous Animals (CEVAP), São Paulo State University (UNESP - Univ Estadual Paulista), Botucatu, SP, Brazil.

 

ABSTRACT

Background

Peripheral 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.

 

Methods

Five 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.

 

Results

The 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.

 

Conclusions

Overall, 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.

 

Keywords Neurorrhaphy; Sciatic nerve; Human embryonic stem cells; Fibrin sealant; FGF2

 

Received: November 08, 2017.

Accepted: March 16, 2018.

 

Correspondence: alroliv@unicamp.br

 

Authors’ contributions

RM, SK, MVC, and ALRO conceived the work, performed experiments, interpreted the results and wrote the manuscript. RSF Jr and BB provided the heterologous fibrin sealant and contributed to the revision of the manuscript. All authors read and approved the final manuscript.

 

Competing interests

The authors declare that they have no competing interests.